Lipidomic biomarkers for identification of high-risk coronary artery disease patients

ABSTRACT

The present invention inter alia provides a method, and use thereof, of predicting severe CVD complications such as AMI or CVD death by detecting the lipid concentrations or lipid ratios of a biological sample and comparing it to a control and has identified specific lipid markers that are more specific and sensitive in predicting these CVD complications than currently utilized clinical markers. Also provided is an antibodies towards said lipids, and the use thereof for predicting, diagnosing, preventing and/or treating CVD complications. The invention additionally relates to kits comprising lipids and/or an antibody thereto, for use in the prediction and/or diagnosis of CVD complications.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No. 14/875,087 filed on 5 Oct. 2015 (now U.S. Pat. No. 9,841,431), which is a continuation of U.S. application Ser. No. 13/805,319, filed 18 Dec. 2012 with a 371 date of 13 Feb. 2013 (now U.S. Pat. No. 9,201,080), which is a U.S. National Stage application of PCT/EP2011/060253 filed 20 Jun. 2011, which claims the benefit of priority of European patent application 10006399.9 filed 20 Jun. 2010 and U.S. Provisional Patent Application 61/356,675 filed 21 Jun. 2010, the entire disclosures of which are herein incorporated by reference in their entireties.

FIELD OF THE INVENTION

This invention relates to methods and uses involving lipid levels to predict and prevent severe cardiovascular disease-associated fatal complications. The invention thus provides a means to identify and treat high-risk coronary artery disease patients. The methods include analyzing lipid levels of a biological sample, and comparing it to a control.

BACKGROUND OF THE INVENTION

Worldwide, cardiovascular diseases (CVD) are among the leading causes of mortality and morbidity with ever-increasing prevalence. CVD is used to classify numerous conditions that affect the heart, heart valves, blood, and vasculature of the body. One of these conditions is coronary artery disease (CAD). Early targeted initiation of preventive measures of CVD-related fatal complications, such as acute myocardial infarction (AMI) and death, would be of great benefit and can provide a major opportunity in reducing mortality and morbidity in patients suffering from CVD. To this end, accurate identification of individuals who are at risk of developing CVD complications is essential. However, traditional risk assessment fails to recognize a substantial proportion of patients at high risk while a large proportion of individuals are classified as having intermediate risk, leaving patient management uncertain. Additional strategies to further refine risk assessment of high-risk CVD are therefore highly needed. To this end, the inventors have evaluated the role of novel lipidomic biomarkers as a prognostic tool for fatal cardiovascular events in CVD patients.

Plasma or serum total cholesterol, LDL-cholesterol or HDL-cholesterol concentrations have been used as gold standard biomarkers for CVD/CAD risk prediction. However, a number of coronary artery disease (CAD) or acute myocardial infarction (AMI) patients have LDL-C levels within the recommended range suggesting the need for additional diagnostic measures of the residual risk. It is evident from earlier large scale population studies that these measurements associate with the CAD risk and CAD endpoints such as AMI or cardiovascular death. Therefore, preventive treatment strategies have so far been addressed to lower LDL-C concentrations (mainly by statin treatment) and more recently also attempts to raise HDL-C have been made (e.g., by CETP-inhibitors). On the other hand, it has also been observed that one half of the AMI patients actually do have normal LDL cholesterol levels and that there is a substantial residual risk in statin treated patients despite a LDL-C lowering. Furthermore, recent publications have demonstrated that plasma levels of apolipoprotein B (apoB), the main surface protein on LDL particles, and LDL-C, the amount of cholesterol in those particles, are correlated and, considered separately, as positive risk factors. Plasma levels of apolipoprotein A₁, the main surface protein on HDL particles, and HDL-C, the amount of cholesterol in those particles, are also correlated with each other and, considered separately, as negative risk factors. Importantly, for a given usual apoB, lower LDL-C has been observed to associate with a higher risk of AMI supporting the view that, on average, LDL particles with low cholesterol content per particle (small, dense LDL particles) are particularly hazardous. Thus, it seems possible that LDL-C associates directly with the more dangerous molecules carried by LDL-particle and that LDL-C is only an indirect measurement of the risk. Therefore, it is of importance to search for molecules e.g., certain lipid species that are directly related with hazardous (i.e., fatal) cardiovascular events.

Lipid metabolite imbalance is a probable cause of dyslipidemia and the ensuing atherosclerosis manifested in its gravest form as the vulnerable atherosclerotic plaque. Atherosclerotic plaques are complex molecular formations that contain numerous lipids. However, there are other factors than lipid rich plaques or LDL cholesterol that make lipids an attractive group of molecules for CVD studies. Lipids are tightly regulated which makes Lipidomic data robust and informative on the current state of the studied organism. Also, lipids are one of the culmination points of a biological system, more the true outcome than the predictor. Combining Lipidomic data with appropriate biobanked clinical material presents a good opportunity for biomarker discovery. Moreover, lipidomics can be used as a gauge of efficacy and safety in drug development and evolving theragnostics. Lipidomic biomarkers are prime candidates for true companion diagnostics in the CVD area and present many opportunities for improved translational medicine as well.

The plaque building blocks and lipoprotein components that are thought to traffic lipids to the site of lesion formation can now be resolved with Lipidomic studies correlating lipid structure and composition to function and thereby disease pathogenesis. While the number of lipid mediators in the human body is overwhelming, their identification and quantification is facilitated by the advances in mass spectrometry and lipid biochemistry, which today enable the simultaneous high throughput identification and quantification of hundreds of molecular lipid species in several lipid classes (Ejsing C S, et al: Global analysis of the yeast lipidome by quantitative shotgun mass spectrometry. Proc Natl Acad Sci USA 2009, 106:2136-2141; Stahlman M, et al: High-throughput shotgun lipidomics by quadrupole time-of-flight mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci 2009 Hiukka A, et al: ApoCIII-enriched LDL in type 2 diabetes displays altered lipid composition, increased susceptibility for sphingomyelinase, and increased binding to biglycan. Diabetes 2009, 58:2018-2026; Linden D, et al: Liver-directed overexpression of mitochondrial glycerol-3-phosphate acyltransferase results in hepatic steatosis, increased triacylglycerol secretion and reduced fatty acid oxidation. FASEB J 2006, 20:434-443.) collectively referred to as the lipidome. Lipidomic studies identify lipid cellular distribution and describe their biochemical mechanisms, interactions and dynamics. Importantly, lipidomics quantifies the exact chemical composition of lipidomes (Han X, Gross R W: Global analyses of cellular lipidomes directly from crude extracts of biological samples by ESI mass spectrometry: a bridge to lipidomics. J Lipid Res 2003, 44:1071-1079).

Due to both high sensitivity and selectivity of lipidomics, even the smallest sample amounts can be analyzed today. The bulk of the lipid data in the art today presents lipids in a sum composition format, i.e. phosphatidylcholine (PC) 34:1 (Brugger B, et al: Quantitative analysis of biological membrane lipids at the low picomole level by nano-electrospray ionization tandem mass spectrometry. Proc Natl Acad Sci USA 1997, 94:2339-2344) where the molecular lipid and the attached fatty acid tails remain unidentified. The identification of molecular lipid species, e.g., PC 16:0/18:1 (Ekroos K, et al: Charting molecular composition of phosphatidylcholines by fatty acid scanning and ion trap MS3 fragmentation. J Lipid Res 2003, 44:2181-2192) is the main feature of advanced lipidomics, which delivers highly resolved molecular lipid species rather than summed fatty acid information. For example, the information of the type of fatty acids and their positions of attachment to the glycerol backbone making up the particular PC molecule is revealed. There are conventional techniques such as thin-layer chromatography combined with gas chromatography but they not only require considerably larger sample amounts and laborious sample preparation, but they do not deliver the molecular lipid species. Despite multiple mass spectrometry techniques capable of characterizing lipid entities, most of them are still unable to deliver reliable high-quality quantitative data in terms of absolute or close-to absolute concentrations. In the context of the present invention, electrospray ionization mass spectrometry-based lipidomics is the preferred technology and can utilize both shotgun and targeted lipidomics for exhaustive deciphering and precise quantification of molecular lipidomes. The superior quality and specificity of shotgun and targeted lipidomics will meet stringent regulatory standards, such as good laboratory practice guidelines (GLP) when set-up in the proper environment. Using these technologies quantification of up to two thousand molecular lipids is possible even in a high throughput format.

Lipidomics is a tool for differentiating patients based on their molecular lipid profiles. Personalized medicine and diagnostics enabled by lipidomics will facilitate the mission of the right individual receiving the right drug at the right time and dose. Several works employing analytes consisting of lipids, proteins and hydrophilic molecules among many others have been conducted to meet the needs of personalized medicine. Recently, non-hypothesis-driven metabolomic screenings have been used to identify novel CVD biomarkers.

For example, WO2004/038381 discloses a method for metabolomically facilitating the diagnosis of a disease state of a subject, or for predicting whether a subject is predisposed to having a disease state wherein the small molecule profile from a subject is obtained and compared to a standard small molecule profile.

WO2008/148857 discloses a method to assess the risk of cardiovascular disease in a patient (including atherosclerosis) by isolating the HDL fraction and sub-fraction from a blood sample of the patient. The components of the HDL fraction or sub-fraction to be measured were Sphingosine-1-Phosphate (S1P), sphingomyelin (SM) and Apolipoprotein A-I (apoA-1).

WO2008/11943 further discloses markers for detecting coronary artery disease that can indicate a patient at risk of having or developing coronary artery disease. These include “first-choice” molecules which were: C18:3 Cholesterol ester, C32:1 Phosphatidylcholine, Alanine, Lipid (mainly VLDL), Lysine, Hexadecanoic acid, C36:2 Phosphatidylcholine, Formate, C32:2 Phosphatidylcholine, C18:2 (Linoleic Acid), Cholesterol, C 18:2 Lyso-phosphatidylcholine, C36:3 Phosphatidylcholine, C34:4 Phosphatidylcholine and C34:3 Phosphatidylcholine.

Furthermore, US2007/0099242 describes a method to determine if a subject is at risk to develop, or is suffering from cardiovascular disease. The method involves determining a change in the amount of a biomarker in the biological sample or HDL sub-fraction thereof, compared to a control sample, wherein the biomarker is at least one of Apolipoprotein C-IV (“ApoC-IV”), Paraoxonase 1 (“PON-1”), Complement Factor 3 (“C3”), Apolipoprotein A-IV (“ApoA-IV”), Apolipoprotein E (“ApoE”), Apolipoprotein LI (“ApoLl”), Complement Factor C4 (“C4”), Complement Factor C4B1 (“C4B1”), Histone H2A, Apolipoprotein C-II (“ApoC-II”), Apolipoprotein M (“ApoM”), Vitronectin, Haptoglobin-related Protein and Clusterin. The document also discloses a method for detecting the presence of one or more atherosclerotic lesions wherein a change in the amount of a biomarker in the biological sample or HDL sub-fraction thereof is detected, compared to a control sample and wherein the biomarker is selected from PON-1, C3, C4, ApoE, ApoM and C4B1. All biomarkers mentioned in this document are protein or lipoprotein biomarkers.

From previous work it cannot be extrapolated that lipid analysis will yield by default a CVD biomarker predictive to the fatal outcomes associated with CVD/CAD. The present invention identifies biomarkers of high risk CVD by absolute, or close to absolute, quantification of defined molecular lipid species instead of profiling multiple analytes. Importantly, while many of the existing biomarker candidates are composite fingerprints of multiple factors, the lipidomics approach herein shows value already at a level of single species or ratios thereof.

In the present invention herein, lipid biomarker concentrations have been measured and quantified in patients with documented CAD who did not show fatal outcomes during the follow-up period (3 years) and in high-risk CAD patients who died due to cardiovascular events during the follow-up period. This invention thus enables accurate usage of the lipid-based biomarkers to identify high risk CVD/CAD patients. Another layer of accuracy was reached through a careful patient selection since it is important to control for factors which may affect the lipid concentration read-outs. Unlike the previous efforts described above, we used specific targeted platforms on a singular technology set-up to analyze lipid species in particular.

The technology and the way it was applied in the context of the inventive teaching presented herein is set apart from similar efforts in the field inter alia due to the following criteria. In sample preparation, samples are strictly controlled and treated identically to avoid potential artifacts that could arise from improper handling. In connection with the present invention, samples were carefully thawed slowly on ice and directly thereafter subjected to a custom-made automated lipid extraction which possesses currently the highest precision in liquid handling, therefore minimizing potential errors. Furthermore, sample freeze-thaw cycles were strictly controlled since this can dramatically affect the lipid stabilities. The automated lipid extraction is based on the method by Folch and colleagues (Folch J, et al: A simple method for the isolation and purification of total lipids from animal tissues. J Biol Chem 1957, 226(1):497-509) which uses chloroform and methanol. This method is preferred when a wide range, from polar to non-polar, of lipid classes are to be extracted with optimal recoveries thus preventing the loss of lipid species. Lipid class specific non-endogenous lipids, when applicable, were used as internal standards to gain highest precision in identification (minimizing false positives) and quantification of monitored molecular lipid species. In this way absolute or semi-absolute amounts of endogenous molecular lipids were determined with the highest precision that can be achieved with today's technologies. The endogenous lipids and respective standards were monitored at the molecular lipid level. In this way, not only false positive identifications were minimized, but molecular lipids could be precisely determined and quantified. Analysis quality was strictly controlled using a novel quality control system. This was mainly controlled by multiple internal standards (IS), external standards (ES), IS/ES ratios, and instrument control samples. By stringently controlling these components, technical and biological outliers were readily identified and rejected from further analysis. To obtain best precision in sensitivity, selectivity and quantification for each molecular lipid different targeted platforms were used. Some lipids are best analyzed using high performance liquid chromatography (HPLC) or ultra high performance liquid chromatography (UHPLC) combined with mass spectrometry based multiple reaction monitoring (MRM) whereas others are best analyzed by direct infusion in combination with mass spectrometry-based precursor ion scanning and neutral loss scanning techniques.

SUMMARY OF THE INVENTION

The present invention provides novel lipidomic markers for predicting and preventing severe CVD/CAD-associated complications, including AMI and death. These markers thus provide a means to identify and treat high-risk coronary artery disease patients. Specifically, it has been found that the lipid molecules, lipid-lipid ratios and lipid-clinical concentration ratios provided herein, when displaying an increased or decreased level—as the case may be—in samples from CAD patients, are useful lipidomic markers for the methods and uses in accordance with the present invention. These sensitive and specific markers were specifically tested to display superior diagnostic and prognostic value compared to the current clinically-used markers predictive for CVD/CAD outcomes. In fact, the currently available biomarkers such as LDL-C or HDL-C have only very limited or no value in predicting the CVD death risk in CAD patients. The present invention therefore represents a significant advantage to other markers which are currently used to diagnose and/or predict CVD and CVD complications, which include LDL-C, total plasma/serum cholesterol and Apolipoprotein B and A1. Thus, the lipidomic markers provided herein allow better diagnosis of or assessment of the risk to develop major CVD complications such as AMI or CVD death.

In accordance with the present invention, methods are inter alia disclosed herein for determining the risk of a patient to develop CVD complications, determining warning signs of CVD risks, (including death, myocardial infarction (MI), angina pectoris, transischemic attack (TIA) and stroke) in said patient.

Methods according to the invention typically comprise the steps of: a) providing a biological sample from a CAD subject; b) determining a lipid concentration, lipid-lipid ratio, or lipid-clinical concentration ratio or (a) corresponding profile(s) from said sample (i.e., determining information on a lipidomic marker in accordance with the invention); and c) comparing said determined lipid concentration, lipid-lipid ratio, or lipid-clinical concentration ratio or said corresponding profile(s) to the corresponding lipid concentration, lipid-lipid ratio, or lipid-clinical concentration ratio or the corresponding profile(s) in a control.

The control may be a sample from (a) CAD patient(s) with no history of major CVD events. It may also be a sample that represents a combination of samples from a CAD patient population with no history of major CVD events. Alternatively, the control may be a set of data concerning a lipidomic marker in accordance with the present invention, e.g., information on the concentration of (a) lipid(s), lipid-lipid ratio(s), or lipid-clinical concentration ratio(s) in accordance with the present invention in a sample when taken from (a) CAD patient(s) with no history of major CVD events, or in a combination of samples taken from a CAD patient population with no history of major CVD events. Said information, and thus the corresponding set of data, may have been previously determined, calculated or extrapolated, or may have yet to be determined, calculated or extrapolated, or may also be taken from the literature.

As mentioned above, the lipidomic marker to be compared between the subject sample and the control (or control sample) may be one or more of the lipid concentration(s), lipid-lipid ratio(s), or lipid-clinical concentration ratio(s) or combinations thereof, i.e., the corresponding profile(s), as described and claimed herein. In this regard, the control or control sample allows establishment of the lipidomic marker baseline or starting point.

In connection with all aspects and embodiments of the invention described and claimed herein, the determination of the lipid concentration(s), the lipid-lipid ratio(s) or the lipid-clinical concentration ratio(s) is typically performed using an assay. Collecting information on a lipidomic marker (i.e., the concentration(s) of (a) lipid(s), lipid-lipid ratio(s), or lipid-clinical concentration ratio(s) or combinations thereof, i.e., corresponding profile(s)) from the sample of a patient and, where appropriate, a corresponding control sample, can be performed with various chemical and high-resolution analytical techniques. Suitable analytical techniques include, but are not limited to, mass spectrometry and nuclear resonance spectroscopy. Any high-resolution technique capable of resolving individual lipids or lipid classes and providing structural information of the same can be used to collect the information on the lipidomic marker in question, e.g., lipid profile from the biological sample. Collecting the information on the lipidomic marker with mass spectrometry (MS) is one of the preferred embodiments of the current invention. The MS instrument can be coupled to a direct sample infusion method, such as a robotic nanoflow ion source device, or to a high performance separation method such as high performance liquid chromatography (HPLC) or ultra performance liquid chromatography (UPLC).

Again in accordance with all aspects and embodiments described and claimed herein, both the sample from the subject and the control sample is preferably a blood sample, more preferably a blood plasma sample, or also preferably a blood serum sample. It may also be a fraction of blood, blood plasma or blood serum, e.g., a lipoprotein fraction. A blood sample can be prepared and plasma or serum, or fractions thereof, can be separated therefrom with techniques well known to the person skilled in the art. Alternatively, both the sample from the subject and the control sample may also be a tissue sample, e.g., artery tissue, such as carotid artery tissue, or artery plaque material, such as carotid artery plaque material.

The lipidomic markers of the present invention allow for prediction and prevention of fatal CVD complications. This will facilitate earlier intervention, less symptom development and suffering and decreased morbidity/mortality associated with CVD.

Thus, the lipidomic markers described and claimed herein allow for individual tailoring of drug intervention for patients being at risk to develop major CVD complications.

In other words, the present invention discloses diagnostic and/or predictive lipid markers and lipid-lipid or lipid-clinical concentration ratios for use in predicting CVD complications such as AMI or CVD death. The invention uses the measurement of lipid concentrations, lipid-lipid and/or lipid-clinical concentration ratios to determine the risk of said subject to develop CVD complications such as AMI and/or CVD death. The subject may have previously suffered from a cardiovascular disease event such as angina pectoris, myocardial infarction or stroke. The CVD may or may not be a result of atherosclerosis.

Accordingly, in one aspect of the invention, a method is provided for determining whether a subject is at risk to develop one or more CVD complications, such as AMI or CVD death, said method comprising determining in a sample from said subject the concentration(s) of one or more lipid(s), wherein (an) increased or decreased concentration(s) in said sample, when compared to a control sample, is (are) indicative of said subject having an increased risk of developing one or more CVD complications, such as AMI or CVD death, wherein the one or more lipid(s) whose increase(s) in concentration is (are) compared to the control is (are) selected from (Tables 4a and 7a): Cer(d18:1/18:0), Cer(d18:1/20:0), Cer(d18:1/24:1), GlcCer(d18:1/18:0), LacCer(d18:1/18:0), LacCer(d18:1/20:0), LacCer(d18:1/22:0), LacCer(d18:1/24:0), PC O-32:0 (KDdiA-PC), PS O-16:0/18:2-alkenyl, PS O-16:1/18:2-alkyl, PS O-18:0/18:2-alkenyl (PS O-18:1/18:2-alkyl), PS O-18:2/16:0-alkenyl and Total LacCer; and wherein the one or more lipid(s) whose decrease(s) in concentration is (are) compared to the control is (are) selected from (Tables 4a and 7a):

CE 14:0, CE 16:0, CE 17:1, CE 20:3, Cer(d18:0/22:0), Cer(d18:0/24:0), LPC 18:1, PC 16:0/18:2, PC 16:0/20:3, PC 16:0/20:4, PC 16:0/22:6, PC 18:0/18:1, PC 18:0/20:3, PC 18:0/20:4, PC 18:1/18:2, SM (d18:1/14:0) (d18:1/13:1-OH), SM (d18:1/23:0) (d18:1/22:1-OH), SM (d18:1/24:0) (d18:1/23:1-OH), Total CE, Total LPC and Total PC.

In a particular embodiment, a method is provided for determining whether a subject is at risk to develop one or more CVD complications, such as AMI or CVD death, wherein the subject is not undergoing statin treatment and wherein said method comprises determining in a sample from said subject the concentration(s) of one or more lipid(s), wherein (an) increased or decreased concentration(s) in said sample, when compared to a control sample, is (are) indicative of said subject having an increased risk of developing one or more CVD complications, such as AMI or CVD death, and wherein the one or more lipid(s) whose increase in concentration is (are) compared to the control is (are) selected from (Tables 4b and 7b):

Cer(d18:1/16:0), Cer(d18:1/18:0), Cer(d18:1/20:0), Cer(d18:1/22:0), Cer(d18:1/24:1), Cer(d18:1/26:1), GlcCer(d18:1/18:0), GlcCer(d18:1/20:0), GlcCer(d18:1/24:1), GlcCer(d18:1/26:1), LacCer(d18:1/18:0), LacCer(d18:1/20:0), LacCer(d18:1/22:0), LacCer(d18:1/24:0), LacCer(d18:1/24:1), PC O-32:0 (KDdiA-PC), PS O-16:0/18:2-alkenyl, PS O-16:1/18:2-alkyl, PS O-18:2/16:0-alkenyl, Total Cer, Total DAG and Total LacCer;

and wherein the one or more lipid(s) whose decrease(s) in concentration is (are) compared to the control is (are) selected from (Tables 4b and 7b):

CE 14:0, CE 17:1, CE 20:3, Cer(d18:0/24:0), LPC 18:1, PC 16:0/20:3, PC 16:0/20:4, PC 18:0/20:4, PC O-40:3, SM (d18:1/14:0) (d18:1/13:1-OH), Total LPC and Total PC.

In a preferred embodiment (for subjects that are either undergoing or not undergoing statin treatment), the one or more lipid(s) whose increase(s) in concentration is (are) compared to the control is (are) selected from (Table 8):

Cer(d18:1/20:0), LacCer(d18:1/20:0), Cer(d18:1/24:1), LacCer(d18:1/24:1), PS O-18:2/16:0-alkenyl, PS O-16:1/18:2-alkyl, Total Cer, Total LacCer, GlcCer(d18:1/24:1), LacCer(d18:1/22:0) and Cer(d18:1/18:0).

In another preferred embodiment, the one or more lipid(s) whose decrease(s) in concentration is (are) compared to the control is (are) selected from (Table 8):

Total PC, PC 16:0/20:4, Cer(d18:0/24:0), Total LPC, CE 14:0, CE 20:3, CE 17:1, PC 16:0/20:3, LPC 18:1, PC 18:0/20:3, PC 18:0/18:1 and Cer(d18:0/22:0).

In a particularly preferred embodiment, the one or more lipid(s) whose increase(s) in concentration is (are) compared to the control is (are) selected from (Table 11):

Cer(d18:1/20:0), LacCer(d18:1/20:0), Cer(d18:1/24:1), LacCer(d18:1/24:1), LacCer(d18:1/22:0) and Cer(d18:1/18:0).

In another particularly preferred embodiment, the one or more lipid(s) whose decrease(s) in concentration is (are) compared to the control is (are) selected from (Table 11):

PC 16:0/20:4 and Cer(d18:0/24:0).

In an alternative embodiment, the present invention relates to a method for determining whether a subject is at risk to develop one or more complications such as AMI or CVD death, comprising determining in a sample from said subject one or more lipid-lipid ratio(s), wherein (an) increased or decreased lipid-lipid ratio(s) in said sample, when compared to a control sample, is (are) indicative of said subject having an increased risk of developing one or more CVD complications, such as AMI or CVD death, wherein the one or more lipid-lipid ratio(s) whose increase(s) is (are) compared to the control is (are) selected from (Tables 5a and 8a):

CE 16:0/CE 18:3, CE 18:2/CE 18:3, CE 19:1/Cer(d18:0/22:0), Cer(d18:1/16:0)/LPC 18:1, Cer(d18:1/16:0)/Total PC, Cer(d18:1/18:0)/LPC 16:0, Cer(d18:1/18:0)/LPC 18:1, Cer(d18:1/18:0)/PC 16:0/18:1, Cer(d18:1/18:0)/PC 16:0/20:3, Cer(d18:1/18:0)/PC 16:0/20:4, Cer(d18:1/18:0)/PC 18:0/20:4, Cer(d18:1/18:0)/Total LPC, Cer(d18:1/18:0)/Total PC, Cer(d18:1/20:0)/PC 16:0/18:1, Cer(d18:1/20:0)/PC 16:0/20:3, Cer(d18:1/20:0)/PC 16:0/20:4, Cer(d18:1/20:0)/PC 18:0/20:4, Cer(d18:1/20:0)/PC 18:1/18:1, Cer(d18:1/20:0)/Total PC, Cer(d18:1/22:0)/LPC 18:2, Cer(d18:1/22:0)/PC 16:0/20:3, Cer(d18:1/22:0)/PC 16:0/20:4, Cer(d18:1/22:0)/Total PC, Cer(d18:1/24:1)/LPC 18:1, Cer(d18:1/24:1)/LPC 18:2, Cer(d18:1/24:1)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 16:0/20:4, Cer(d18:1/24:1)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/20:3, Cer(d18:1/24:1)/PC 18:0/20:4, Cer(d18:1/24:1)/SM (d18:1/24:0) (d18:1/23:1-OH), Cer(d18:1/24:1)/Total CE, Cer(d18:1/24:1)/Total LPC, Cer(d18:1/24:1)/Total PC, Cer(d18:1/26:0)/PC O-40:0, GlcCer(d18:1/20:0)/PC 16:0/20:4, GlcCer(d18:1/20:0)/Total PC, GlcCer(d18:1/26:0)/Total CE, LacCer(d18:1/16:0)/Total LPC, LacCer(d18:1/18:0)/PC 16:0/18:1, LacCer(d18:1/18:0)/PC 16:0/20:3, LacCer(d18:1/18:0)/PC 18:0/18:1, LacCer(d18:1/18:0)/PC 18:0/20:3, LacCer(d18:1/18:0)/PC 18:1/18:1, LacCer(d18:1/18:0)/PC 18:1/18:2, LacCer(d18:1/18:0)/Total LPC, LacCer(d18:1/18:0)/Total PC, LacCer(d18:1/20:0)/PC 16:0/18:1, LacCer(d18:1/20:0)/PC 16:0/20:3, LacCer(d18:1/20:0)/PC 18:0/18:1, LacCer(d18:1/20:0)/PC 18:0/20:3 LacCer(d18:1/20:0)/PC 18:1/18:1, LacCer(d18:1/20:0)/PC 18:1/18:2, LacCer(d18:1/20:0)/PC 18:2/18:2, LacCer(d18:1/20:0)/SM (d18:1/17:1-OH), LacCer(d18:1/20:0)/SM (d18:1/18:0), LacCer(d18:1/20:0)/Total LPC, LacCer(d18:1/20:0)/Total PC, LacCer(d18:1/20:0)/Total SM, LacCer(d18:1/22:0)/PC 16:0/20:3, LacCer(d18:1/22:0)/PC 16:0/20:4, LacCer(d18:1/22:0)/PC 18:0/20:3, LacCer(d18:1/22:0)/SM (d18:1/14:0) (d18:1/13:1-OH), LacCer(d18:1/22:0)/Total LPC, LacCer(d18:1/22:0)/Total PC, LacCer(d18:1/24:0)/PC 16:0/20:3, LacCer(d18:1/24:0)/Total LPC, LacCer(d18:1/24:1)/Total LPC, LacCer(d18:1/24:1)/Total PC, LacCer(d18:1/24:1)/Total PC O, PC 16:0/16:0/PC 16:0/20:4, PC 16:0/16:0/Total PC, PC 16:0/18:2/Total PC, PC O-18:0/18:2-alkyl/PC O-36:5, PC O-32:0 (KDdiA-PC)/PC O-38:5, PS O-16:0/18:2-alkenyl/Total PS O, PS O-16:1/18:2-alkyl/Total PS O, PS O-18:0/18:2-alkenyl (PS O-18:1/18:2-alkyl)/Total PS O, PS O-18:2/16:0-alkenyl/Total PS O, Total Cer/Total PC, Total LacCer/Total PC and Total LacCer/Total PC O;

and wherein the one or more lipid-lipid ratio(s) whose decrease(s) is (are) compared to the control is (are) selected from (Tables 5a and 8a):

CE 14:0/Cer(d18:1/24:1), CE 16:0/Cer(d18:1/24:1), CE 16:1/CE 19:1, CE 16:1/Cer(d18:1/18:0), CE 16:1/Cer(d18:1/20:0), CE 16:1/Cer(d18:1/24:1), CE 16:1/GlcCer(d18:1/18:0), CE 16:1/GlcCer(d18:1/20:0), CE 16:1/LacCer(d18:1/16:0), CE 16:1/LacCer(d18:1/18:0), CE 16:1/LacCer(d18:1/20:0), CE 16:1/LacCer(d18:1/22:0), CE 16:1/LacCer(d18:1/24:0), CE 16:1/PC 16:0/16:0, CE 16:1/Total LacCer, CE 17:1/Cer(d18:1/24:1), CE 17:1/GlcCer(d18:1/24:1), CE 17:1/LacCer(d18:1/18:0), CE 18:1/Total LacCer, CE 18:3/Cer(d18:1/16:0), CE 18:3/Cer(d18:1/18:0), CE 18:3/Cer(d18:1/20:0), CE 18:3/Cer(d18:1/22:0), CE 18:3/Cer(d18:1/24:0), CE 18:3/Cer(d18:1/24:1), CE 18:3/GlcCer(d18:1/18:0), CE 18:3/GlcCer(d18:1/20:0), CE 18:3/LacCer(d18:1/18:0), CE 18:3/LacCer(d18:1/20:0), CE 18:3/LacCer(d18:1/22:0), CE 18:3/LacCer(d18:1/24:0), CE 18:3/PC 16:0/16:0, CE 18:3/PC O-34:1, CE 18:3/PS O-16:0/18:1-alkenyl (PS O-16:1/18:1-alkyl), CE 18:3/PS O-16:0/18:2-alkenyl, CE 18:3/PS O-16:1/18:2-alkyl, CE 18:3/Total CE, CE 18:3/Total Cer, CE 18:3/Total LacCer, CE 20:3/Cer(d18:1/24:1), CE 20:3/LacCer(d18:1/20:0), Cer(d18:0/22:0)/Cer(d18:1/18:0), Cer(d18:0/22:0)/Cer(d18:1/20:0), Cer(d18:0/22:0)/Cer(d18:1/22:0), Cer(d18:0/22:0)/Cer(d18:1/24:1), Cer(d18:0/22:0)/LacCer(d18:1/24:0), Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/22:0)/Total CE, Cer(d18:0/24:0)/Cer(d18:1/16:0), Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/Cer(d18:1/20:0), Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/GlcCer(d18:1/20:0), Cer(d18:0/24:0)/LacCer(d18:1/24:0), Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/24:0)/SM (d18:1/17:0) (d18:1/16:1-OH), Cer(d18:0/24:0)/Total CE, Cer(d18:0/24:0)/Total Cer, Cer(d18:0/24:1)/Total CE, GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), GlcCer(d18:1/26:0)/LacCer(d18:1/22:0), LPC 16:0/LacCer(d18:1/20:0), LPC 16:0/LacCer(d18:1/22:0), LPC 16:0/LacCer(d18:1/24:1), LPC 16:0/Total LacCer, LPC 18:1/LacCer(d18:1/20:0), LPC 18:2/LacCer(d18:1/20:0), LPC 18:2/PS O-16:0/18:2-alkenyl, LPC 18:2/PS O-16:1/18:2-alkyl, PC 16:0/20:3/PS O-16:0/18:2-alkenyl, PC 16:0/20:3/PS O-16:1/18:2-alkyl, PC 18:1/18:2/PS O-16:0/18:2-alkenyl, PC 18:1/18:2/PS O-16:1/18:2-alkyl, SM (d18:1/24:0) (d18:1/23:1-OH)/Total Cer and Total LPC/Total LacCer.

In an alternative embodiment, the present invention relates to a method for determining whether a subject is at risk to develop one or more complications such as AMI or CVD death, wherein the subject is not undergoing statin treatment and wherein said method comprises determining in a sample from said subject one or more lipid-lipid ratio(s), wherein (an) increased or decreased lipid-lipid ratio(s) in said sample, when compared to a control sample, is (are) indicative of said subject having an increased risk of developing one or more CVD complications, such as AMI or CVD death, wherein the one or more lipid-lipid ratio(s) whose increase(s) is (are) compared to the control is (are) selected from (Tables 5b and 8b):

CE 16:0/CE 18:3, CE 18:0/CE 18:3, CE 18:2/CE 18:3, Cer(d18:1/16:0)/LPC 18:1, Cer(d18:1/16:0)/Total PC, Cer(d18:1/18:0)/LPC 18:1, Cer(d18:1/18:0)/PC 16:0/18:1, Cer(d18:1/18:0)/PC 16:0/20:3, Cer(d18:1/18:0)/PC 16:0/20:4, Cer(d18:1/18:0)/PC 18:0/18:1, Cer(d18:1/18:0)/PC 18:0/20:4, Cer(d18:1/18:0)/PC 18:1/18:1, Cer(d18:1/18:0)/SM (d18:1/14:0) (d18:1/13:1-OH), Cer(d18:1/18:0)/SM (d18:1/17:2-OH), Cer(d18:1/18:0)/SM (d18:1/18:1), Cer(d18:1/18:0)/Total CE, Cer(d18:1/18:0)/Total LPC, Cer(d18:1/18:0)/Total PC, Cer(d18:1/20:0)/PC 16:0/18:1, Cer(d18:1/20:0)/PC 16:0/20:3, Cer(d18:1/20:0)/PC 16:0/20:4, Cer(d18:1/20:0)/PC 18:0/18:1, Cer(d18:1/20:0)/PC 18:0/20:4, Cer(d18:1/20:0)/PC 18:1/18:1, Cer(d18:1/20:0)/SM (d18:1/14:0) (d18:1/13:1-OH), Cer(d18:1/20:0)/Total LPC, Cer(d18:1/20:0)/Total PC, Cer(d18:1/20:0)/Total PC O, Cer(d18:1/22:0)/LPC 18:2, Cer(d18:1/22:0)/PC 16:0/20:3, Cer(d18:1/22:0)/PC 16:0/20:4, Cer(d18:1/22:0)/PC 18:0/20:4, Cer(d18:1/22:0)/Total PC, Cer(d18:1/24:1)/LPC 18:1, Cer(d18:1/24:1)/LPC 18:2, Cer(d18:1/24:1)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:2, Cer(d18:1/24:1)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 16:0/20:4, Cer(d18:1/24:1)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:2, Cer(d18:1/24:1)/PC 18:0/20:3, Cer(d18:1/24:1)/PC 18:0/20:4, Cer(d18:1/24:1)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:2, Cer(d18:1/24:1)/PC O-40:3, Cer(d18:1/24:1)/SM (d18:1/17:1-OH), Cer(d18:1/24:1)/SM (d18:1/18:0), Cer(d18:1/24:1)/SM (d18:1/23:0) (d18:1/22:1-OH), Cer(d18:1/24:1)/SM (d18:1/24:0) (d18:1/23:1-OH), Cer(d18:1/24:1)/Total CE, Cer(d18:1/24:1)/Total LPC, Cer(d18:1/24:1)/Total PC, GlcCer(d18:1/26:0)/Total CE, GlcCer(d18:1/26:1)/Total CE, LacCer(d18:1/18:0)/Total PC, LacCer(d18:1/20:0)/PC 16:0/20:3, LacCer(d18:1/20:0)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 18:0/18:1, LacCer(d18:1/20:0)/PC 18:0/20:3, LacCer(d18:1/20:0)/PC 18:0/20:4, LacCer(d18:1/20:0)/PC 18:1/18:2, LacCer(d18:1/20:0)/SM (d18:1/17:1-OH), LacCer(d18:1/20:0)/SM (d18:1/18:0), LacCer(d18:1/20:0)/Total CE, LacCer(d18:1/20:0)/Total LPC, LacCer(d18:1/20:0)/Total PC, LacCer(d18:1/20:0)/Total SM, LacCer(d18:1/24:0)/Total LPC, PC 16:0/16:0/PC 16:0/20:4, PC 16:0/16:0/Total PC, PC O-32:0 (KDdiA-PC)/Total PC O, PS O-16:0/18:2-alkenyl/Total PC, PS O-16:0/18:2-alkenyl/Total PC O, PS O-16:0/18:2-alkenyl/Total PS O, PS O-16:1/18:2-alkyl/Total PC, PS O-16:1/18:2-alkyl/Total PC O, PS O-16:1/18:2-alkyl/Total PS O, PS O-18:2/16:0-alkenyl/Total PC O, PS O-18:2/16:0-alkenyl/Total PS O, SM (d18:1/17:0) (d18:1/16:1-OH)/Total PC O, Total Cer/Total PC, Total DAG/Total LPC, Total DAG/Total PC, Total DAG/Total PC O and Total LacCer/Total PC;

and wherein the one or more lipid-lipid ratio(s) whose decrease(s) is (are) compared to the control is (are) selected from (Tables 5b and 8b):

CE 14:0/Cer(d18:1/18:0), CE 14:0/Cer(d18:1/24:1), CE 14:0/Total DAG, CE 15:0/Cer(d18:1/20:0), CE 16:0/Cer(d18:1/18:0), CE 16:0/Cer(d18:1/24:1), CE 16:1/CE 19:1, CE 16:1/Cer(d18:1/18:0), CE 16:1/Cer(d18:1/20:0), CE 16:1/Cer(d18:1/24:1), CE 16:1/GlcCer(d18:1/24:1), CE 16:1/LacCer(d18:1/18:0), CE 16:1/LacCer(d18:1/24:0), CE 16:1/Total LacCer, CE 17:1/Cer(d18:1/18:0), CE 17:1/Cer(d18:1/24:1), CE 18:2/Cer(d18:1/20:0), CE 18:2/Cer(d18:1/24:1), CE 18:3/Cer(d18:1/16:0), CE 18:3/Cer(d18:1/18:0), CE 18:3/Cer(d18:1/20:0), CE 18:3/Cer(d18:1/22:0), CE 18:3/Cer(d18:1/24:0), CE 18:3/Cer(d18:1/24:1), CE 18:3/GlcCer(d18:1/20:0), CE 18:3/LacCer(d18:1/20:0), CE 18:3/LacCer(d18:1/22:0), CE 18:3/LacCer(d18:1/24:0), CE 18:3/PC 16:0/16:0, CE 18:3/PC O-34:1, CE 18:3/PS O-16:0/18:2-alkenyl, CE 18:3/PS O-16:1/18:2-alkyl, CE 18:3/Total CE, CE 18:3/Total Cer, CE 18:3/Total DAG, CE 18:3/Total LacCer, CE 20:3/Cer(d18:1/24:1), CE 20:3/LacCer(d18:1/20:0), CE 20:4/Cer(d18:1/18:0), CE 20:4/Cer(d18:1/24:1), CE 20:4/GlcCer(d18:1/20:0), CE 20:4/GlcCer(d18:1/24:1), CE 20:4/LacCer(d18:1/20:0), CE 20:5/LacCer(d18:1/20:0), Cer(d18:0/22:0)/Cer(d18:1/18:0), Cer(d18:0/22:0)/Cer(d18:1/20:0), Cer(d18:0/22:0)/Cer(d18:1/24:1), Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/22:0)/Total CE, Cer(d18:0/22:0)/Total DAG, Cer(d18:0/22:0)/Total GlcCer, Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/Cer(d18:1/20:0), Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/24:0)/Total CE, Cer(d18:0/24:0)/Total Cer, DAG 16:0/18:1/Total DAG, GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), LPC 16:0/LacCer(d18:1/24:0), LPC 18:1/LacCer(d18:1/20:0), LPC 18:2/LacCer(d18:1/20:0), PC 16:0/20:3/PS O-16:0/18:2-alkenyl, PC 16:0/20:3/PS O-16:1/18:2-alkyl, PC 16:0/20:4/PS O-16:0/18:2-alkenyl, PC 16:0/20:4/PS O-16:1/18:2-alkyl, PC 16:0/20:4/Total DAG, PC 18:0/20:3/PS O-16:0/18:2-alkenyl, PC 18:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-18:2/16:0-alkenyl, PC 18:0/20:4/PS O-16:0/18:2-alkenyl, PC 18:0/20:4/PS O-16:1/18:2-alkyl, PC 18:1/18:2/PS O-16:0/18:2-alkenyl, PC 18:1/18:2/PS O-16:1/18:2-alkyl, PC 18:1/18:2/Total Cer, PC O-40:3/PS O-18:2/16:0-alkenyl, SM (d18:1/23:0) (d18:1/22:1-OH)/Total DAG, SM (d18:1/24:0) (d18:1/23:1-OH)/Total Cer, Total CE/Total DAG and Total LPC/Total LacCer.

In a preferred embodiment (for subjects that are either undergoing or not undergoing statin treatment), the one or more lipid-lipid ratio(s) whose increase is (are) compared to the control is (are) selected from (Table 9):

GlcCer(d18:1/26:1)/Total CE, Cer(d18:1/24:1)/Total PC, Cer(d18:1/24:1)/PC 16:0/20:4, Cer(d18:1/20:0)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 16:0/20:3, Total Cer/Total PC, Total LacCer/Total PC, LacCer(d18:1/20:0)/PC 18:1/18:2, PS O-16:0/18:2-alkenyl/Total PS O, Cer(d18:1/18:0)/PC 16:0/20:4, LacCer(d18:1/20:0)/Total LPC and LacCer(d18:1/20:0)/PC 16:0/20:4;

In another preferred embodiment, the one or more lipid-lipid ratio(s) whose decrease is (are) compared to the control is (are) selected from (Table 9):

Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/22:0)/Cer(d18:1/24:1), DAG 16:0/18:1/Total DAG, Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/24:0)/Total CE, Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/Total Cer, Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/LacCer(d18:1/24:0), Cer(d18:0/22:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/22:0)/Cer(d18:1/20:0), Cer(d18:0/22:0)/P S O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl, GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), Total LPC/Total LacCer and GlcCer(d18:1/26:0)/LacCer(d18:1/22:0).

In a particularly preferred embodiment, the one or more lipid-lipid ratio(s) whose increase is (are) compared to the control is (are) selected from (Table 11):

Cer(d18:1/24:1)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 16:0/20:3, PS O-16:0/18:2-alkenyl/Total PS O and Cer(d18:1/18:0)/PC 16:0/20:4;

and the one or more lipid-lipid ratio(s) whose decrease is (are) compared to the control is (are) selected from (Table 11):

GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), DAG 16:0/18:1/Total DAG, Cer(d18:0/24:0)/Total Cer, Total LPC/Total LacCer, GlcCer(d18:1/26:0)/LacCer(d18:1/22:0), Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl and Cer(d18:0/24:0)/LacCer(d18:1/24:0);

In yet another alternative embodiment the present invention relates to a method for determining whether a subject is at risk to develop one or more CVD complications, such as AMI or CVD death, comprising determining in a sample from said subject one or more lipid-clinical concentration ratio(s), wherein (an) increased or decreased lipid-clinical concentration ratio(s) in said sample, when compared to a control sample, is (are) indicative of said subject having an increased risk of developing one or more CVD complications, such as AMI or CVD death, wherein the one or more lipid-clinical concentration ratio(s) whose increase is (are) compared to the control is (are) selected from (Tables 6a and 9a):

Cer(d18:1/16:0)/triglycerides, Cer(d18:1/18:0)/apolipoprotein A-I, Cer(d18:1/18:0)/apolipoprotein B, Cer(d18:1/18:0)/HDL cholesterol, Cer(d18:1/18:0)/total cholesterol, Cer(d18:1/18:0)/total-c/HDL-c, Cer(d18:1/18:0)/triglycerides, Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/20:0)/apolipoprotein B, Cer(d18:1/20:0)/HDL cholesterol, Cer(d18:1/20:0)/total cholesterol, Cer(d18:1/20:0)/triglycerides, Cer(d18:1/22:0)/apolipoprotein B, Cer(d18:1/22:0)/LDL cholesterol, Cer(d18:1/22:0)/triglycerides, Cer(d18:1/24:1)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein B, Cer(d18:1/24:1)/LDL cholesterol, Cer(d18:1/24:1)/total cholesterol, Cer(d18:1/24:1)/total-c/HDL-c, Cer(d18:1/24:1)/triglycerides, GlcCer(d18:1/24:0)/total cholesterol, LacCer(d18:1/16:0)/triglycerides, LacCer(d18:1/18:0)/apolipoprotein A-I, LacCer(d18:1/18:0)/apolipoprotein B, LacCer(d18:1/18:0)/HDL cholesterol, LacCer(d18:1/18:0)/LDL cholesterol, LacCer(d18:1/18:0)/total cholesterol, LacCer(d18:1/18:0)/total-c/HDL-c, LacCer(d18:1/18:0)/triglycerides, LacCer(d18:1/20:0)/apoA1/apoB, LacCer(d18:1/20:0)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein B, LacCer(d18:1/20:0)/HDL cholesterol, LacCer(d18:1/20:0)/LDL cholesterol, LacCer(d18:1/20:0)/LDL-c/HDL-c, LacCer(d18:1/20:0)/total cholesterol, LacCer(d18:1/20:0)/total-c/HDL-c, LacCer(d18:1/20:0)/triglycerides, LacCer(d18:1/22:0)/apoA/apoB, LacCer(d18:1/22:0)/apolipoprotein A-I, LacCer(d18:1/22:0)/apolipoprotein B, LacCer(d18:1/22:0)/HDL cholesterol, LacCer(d18:1/22:0)/LDL cholesterol, LacCer(d18:1/22:0)/total cholesterol, LacCer(d18:1/22:0)/total-c/HDL-c, LacCer(d18:1/22:0)/triglycerides, LacCer(d18:1/24:0)/apoA/apoB, LacCer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/24:1)/apolipoprotein B, LacCer(d18:1/24:1)/total cholesterol, LacCer(d18:1/24:1)/triglycerides, PC O-32:0 (KDdiA-PC)/apolipoprotein A-I, PC O-32:0 (KDdiA-PC)/triglycerides, PC O-34:1/triglycerides, PS O-16:0/18:1-alkenyl (PS O-16:1/18:1-alkyl)/triglycerides, PS O-16:0/18:2-alkenyl/triglycerides, PS O-16:1/18:2-alkyl/triglycerides, PS O-18:2/16:0-alkenyl/HDL cholesterol, PS O-18:2/16:0-alkenyl/triglycerides, Total Cer/apolipoprotein A-I, Total Cer/total cholesterol, Total Cer/triglycerides, Total GlcCer/apolipoprotein B, Total GlcCer/total cholesterol, Total LacCer/apolipoprotein A-I, Total LacCer/apolipoprotein B, Total LacCer/total cholesterol and Total LacCer/triglycerides;

and wherein the one or more lipid-clinical concentration ratio(s) whose decrease is (are) compared to the control is (are) selected from (Tables 6a and 9a):

CE 14:0/apolipoprotein B, CE 14:0/LDL cholesterol, CE 14:0/LDL-c/HDL-c, CE 14:0/total cholesterol, CE 14:0/total-c/HDL-c, CE 16:1/apolipoprotein B, CE 16:1/HDL cholesterol, CE 16:1/LDL cholesterol, CE 16:1/total cholesterol, CE 17:1/LDL-c/HDL-c, CE 18:3/apoA1/apoB, CE 18:3/apolipoprotein A-I, CE 18:3/apolipoprotein B, CE 18:3/HDL cholesterol, CE 18:3/LDL cholesterol, CE 18:3/LDL-c/HDL-c, CE 18:3/total cholesterol, CE 18:3/total-c/HDL-c, CE 20:3/apolipoprotein B, CE 20:3/LDL-c/HDL-c, CE 20:3/total-c/HDL-c, CE 20:5/apolipoprotein B, CE 20:5/HDL cholesterol, CE 20:5/LDL cholesterol, Cer(d18:0/22:0)/apolipoprotein B, Cer(d18:0/22:0)/LDL-c/HDL-c, Cer(d18:0/22:0)/total-c/HDL-c, Cer(d18:0/24:0)/apolipoprotein A-I, Cer(d18:0/24:0)/apolipoprotein B, Cer(d18:0/24:0)/HDL cholesterol, Cer(d18:0/24:0)/LDL cholesterol, Cer(d18:0/24:0)/LDL-c/HDL-c, Cer(d18:0/24:0)/total cholesterol, Cer(d18:0/24:0)/total-c/HDL-c, LPC 18:2/apoA1/apoB, LPC 18:2/apolipoprotein B, LPC 18:2/HDL cholesterol, LPC 18:2/LDL cholesterol, LPC 18:2/LDL-c/HDL-c, LPC 18:2/total cholesterol, PC 16:0/20:3/apolipoprotein B, PC 16:0/20:3/HDL cholesterol, PC 16:0/20:3/LDL-c/HDL-c, PC 16:0/20:3/total-c/HDL-c, PC 16:0/20:4/apolipoprotein A-I, PC 16:0/20:4/apolipoprotein B, PC 16:0/20:4/LDL cholesterol, PC 16:0/20:4/LDL-c/HDL-c, PC 16:0/20:4/total cholesterol, PC 16:0/20:4/total-c/HDL-c, PC 18:0/18:1/LDL-c/HDL-c, PC 18:0/20:3/LDL-c/HDL-c, PC 18:0/20:3/total-c/HDL-c, PC 18:0/20:4/apoA1/apoB, Total LPC/LDL-c/HDL-c, Total LPC/total-c/HDL-c, Total PC/apolipoprotein B, Total PC/LDL-c/HDL-c, Total PC/total cholesterol and Total PC/total-c/HDL-c.

In yet another alternative embodiment the present invention relates to a method for determining whether a subject is at risk to develop one or more CVD complications, such as AMI or CVD death, wherein the subject is not undergoing statin treatment and wherein said method comprises determining in a sample from said subject one or more lipid-clinical concentration ratio(s), wherein (an) increased or decreased lipid-clinical concentration ratio(s) in said sample, when compared to a control sample, is (are) indicative of said subject having an increased risk of developing one or more of CVD complications, such as AMI or CVD death, wherein the one or more lipid-clinical concentration ratio(s) whose increase is (are) compared to the control is (are) selected from (Tables 6b and 9b):

Cer(d18:1/16:0)/apolipoprotein A-I, Cer(d18:1/16:0)/LDL cholesterol, Cer(d18:1/16:0)/triglycerides, Cer(d18:1/18:0)/apolipoprotein A-I, Cer(d18:1/18:0)/apolipoprotein B, Cer(d18:1/18:0)/HDL cholesterol, Cer(d18:1/18:0)/total cholesterol, Cer(d18:1/18:0)/total-c/HDL-c, Cer(d18:1/18:0)/triglycerides, Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/20:0)/apolipoprotein B, Cer(d18:1/20:0)/HDL cholesterol, Cer(d18:1/20:0)/LDL cholesterol, Cer(d18:1/20:0)/total cholesterol, Cer(d18:1/20:0)/total-c/HDL-c, Cer(d18:1/20:0)/triglycerides, Cer(d18:1/22:0)/apolipoprotein A-I, Cer(d18:1/22:0)/apolipoprotein B, Cer(d18:1/22:0)/LDL cholesterol, Cer(d18:1/22:0)/total cholesterol, Cer(d18:1/22:0)/triglycerides, Cer(d18:1/24:0)/apolipoprotein A-I, Cer(d18:1/24:0)/apolipoprotein B, Cer(d18:1/24:0)/LDL cholesterol, Cer(d18:1/24:0)/total cholesterol, Cer(d18:1/24:1)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein B, Cer(d18:1/24:1)/HDL cholesterol, Cer(d18:1/24:1)/LDL cholesterol, Cer(d18:1/24:1)/LDL-c/HDL-c, Cer(d18:1/24:1)/total cholesterol, Cer(d18:1/24:1)/total-c/HDL-c, Cer(d18:1/24:1)/triglycerides, GlcCer(d18:1/20:0)/apolipoprotein B, GlcCer(d18:1/20:0)/total cholesterol, GlcCer(d18:1/24:1)/apolipoprotein B, GlcCer(d18:1/26:1)/apolipoprotein A-I, LacCer(d18:1/16:0)/triglycerides, LacCer(d18:1/18:0)/apolipoprotein A-I, LacCer(d18:1/18:0)/apolipoprotein B, LacCer(d18:1/18:0)/total cholesterol, LacCer(d18:1/20:0)/apoA1/apoB, LacCer(d18:1/20:0)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein B, LacCer(d18:1/20:0)/HDL cholesterol, LacCer(d18:1/20:0)/LDL cholesterol, LacCer(d18:1/20:0)/LDL-c/HDL-c, LacCer(d18:1/20:0)/total cholesterol, LacCer(d18:1/20:0)/total-c/HDL-c, LacCer(d18:1/20:0)/triglycerides, LacCer(d18:1/22:0)/apoA/apoB, LacCer(d18:1/22:0)/apolipoprotein A-I, LacCer(d18:1/22:0)/apolipoprotein B, LacCer(d18:1/22:0)/HDL cholesterol, LacCer(d18:1/22:0)/LDL cholesterol, LacCer(d18:1/22:0)/total cholesterol, LacCer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/24:1)/apolipoprotein B, LacCer(d18:1/24:1)/total cholesterol, LacCer(d18:1/24:1)/triglycerides, PC O-34:1/apolipoprotein B, PS O-16:0/18:2-alkenyl/triglycerides, PS O-16:1/18:2-alkyl/triglycerides, Total Cer/apolipoprotein A-I, Total Cer/apolipoprotein B, Total Cer/LDL cholesterol, Total Cer/total cholesterol, Total DAG/apolipoprotein A-I, Total DAG/triglycerides, Total GlcCer/apolipoprotein B, Total LacCer/apolipoprotein A-I, Total LacCer/apolipoprotein B and Total LacCer/total cholesterol.

and wherein the one or more lipid-clinical concentration ratio(s) whose decrease is (are) compared to the control is (are) selected from (Tables 6b and 9b):

CE 14:0/apoA1/apoB, CE 14:0/apolipoprotein B, CE 14:0/LDL-c/HDL-c, CE 14:0/total-c/HDL-c, CE 16:1/apoA1/apoB, CE 18:3/apoA1/apoB, CE 18:3/apolipoprotein A-I, CE 18:3/apolipoprotein B, CE 18:3/HDL cholesterol, CE 18:3/LDL cholesterol, CE 18:3/LDL-c/HDL-c, CE 18:3/total cholesterol, CE 18:3/total-c/HDL-c, CE 20:5/triglycerides, Cer(d18:0/24:0)/apolipoprotein B, Cer(d18:0/24:0)/total cholesterol, Cer(d18:0/24:0)/total-c/HDL-c, PC 18:0/20:4/apoA1/apoB, PC O-38:6/apolipoprotein A-I, Total LPC/apoA1/apoB and Total PC/apolipoprotein A-I.

In a preferred embodiment (for subjects that are either undergoing or not undergoing statin treatment), the one or more lipid-clinical concentration ratio(s) whose increase is (are) compared to the control is (are) selected from (Table 10):

Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein A-I, Total Cer/apolipoprotein A-I, Total LacCer/apolipoprotein A-I, Cer(d18:1/18:0)/apolipoprotein A-I, LacCer(d18:1/22:0)/apolipoprotein A-I, LacCer(d18:1/20:0)/HDL cholesterol and Cer(d18:1/24:1)/apolipoprotein B.

In another preferred embodiment, the one or more lipid-clinical concentration ratio(s) whose decrease is (are) compared to the control is (are) selected from (Table 10):

Cer(d18:0/24:0)/apolipoprotein B, Cer(d18:0/24:0)/total cholesterol, Cer(d18:0/24:0)/apolipoprotein B, PC 16:0/20:4/apolipoprotein B and Cer(d18:0/24:0)/apolipoprotein A-I.

In a particularly preferred embodiment, the lipid-clinical concentration ratio whose increase is compared to the control is (are) selected from (Table 11):

Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein A-I, Total LacCer/apolipoprotein A-I, LacCer(d18:1/20:0)/HDL cholesterol and Cer(d18:1/18:0)/apolipoprotein A-I.

For the purposes of the invention, and particularly for lipid-clinical concentration ratios, an Apolipoprotein A-I measurement may alternatively be an Apolipoprotein A-II measurement.

In another aspect the present invention relates to a method for evaluating the effectiveness of a treatment of CVD and/or one or more of its complications, such as AMI or CVD death, in a subject, comprising determining in a sample from said subject the concentration(s) of one or more lipid(s), wherein (an) increased or decreased concentration(s) in said sample, when compared to a control sample, is (are) indicative of effectiveness of said treatment, wherein the one or more lipid(s) whose decrease(s) in concentration is (are) compared to the control is (are) selected from (Table 2a and 5a):

Cer(d18:1/18:0), Cer(d18:1/20:0), Cer(d18:1/24:1), GlcCer(d18:1/18:0), LacCer(d18:1/18:0), LacCer(d18:1/20:0), LacCer(d18:1/22:0), LacCer(d18:1/24:0), PC O-32:0 (KDdiA-PC), PS O-16:0/18:2-alkenyl, PS O-16:1/18:2-alkyl, PS O-18:0/18:2-alkenyl (PS O-18:1/18:2-alkyl), PS O-18:2/16:0-alkenyl and Total LacCer;

and wherein the one or more lipid(s) whose increase(s) in concentration is (are) compared to the control is (are) selected from (Table 2a and 5a):

CE 14:0, CE 16:0, CE 17:1, CE 20:3, Cer(d18:0/22:0), Cer(d18:0/24:0), LPC 18:1, PC 16:0/18:2, PC 16:0/20:3, PC 16:0/20:4, PC 16:0/22:6, PC 18:0/18:1, PC 18:0/20:3, PC 18:0/20:4, PC 18:1/18:2, SM (d18:1/14:0) (d18:1/13:1-OH), SM (d18:1/23:0) (d18:1/22:1-OH), SM (d18:1/24:0) (d18:1/23:1-OH), Total CE, Total LPC and Total PC.

In a particular embodiment, the present invention relates to a method for evaluating the effectiveness of a treatment of CVD and/or one or more of its complications, such as AMI or CVD death, in a subject, wherein the subject is not undergoing statin treatment and wherein said method comprises, determining in a sample from said subject the concentration(s) of one or more lipid(s), wherein (an) increased or decreased concentration(s) in said sample, when compared to a control sample, is (are) indicative of effectiveness of said treatment, wherein the one or more lipid(s) whose decrease(s) in concentration is (are) compared to the control is (are) selected from (Table 2b and 5b):

Cer(d18:1/16:0), Cer(d18:1/18:0), Cer(d18:1/20:0), Cer(d18:1/22:0), Cer(d18:1/24:1), Cer(d18:1/26:1), GlcCer(d18:1/18:0), GlcCer(d18:1/20:0), GlcCer(d18:1/24:1), GlcCer(d18:1/26:1), LacCer(d18:1/18:0), LacCer(d18:1/20:0), LacCer(d18:1/22:0), LacCer(d18:1/24:0), LacCer(d18:1/24:1), PC O-32:0 (KDdiA-PC), PS O-16:0/18:2-alkenyl, PS O-16:1/18:2-alkyl, PS O-18:2/16:0-alkenyl, Total Cer, Total DAG and Total LacCer;

and wherein the one or more lipid(s) whose increase(s) in concentration is (are) compared to the control is (are) selected from (Table 2b and 5b): CE 14:0, CE 17:1, CE 20:3, Cer(d18:0/24:0), LPC 18:1, PC 16:0/20:3, PC 16:0/20:4, PC 18:0/20:4, PC O-40:3, SM (d18:1/14:0) (d18:1/13:1-OH), Total LPC and Total PC.

In a preferred embodiment (for subjects that are either undergoing or not undergoing statin treatment), the one or more lipid(s) whose decrease in concentration is (are) compared to the control is (are) selected from (Table 8):

Cer(d18:1/20:0), LacCer(d18:1/20:0), Cer(d18:1/24:1), LacCer(d18:1/24:1), PS O-18:2/16:0-alkenyl, PS O-16:1/18:2-alkyl, Total Cer, Total LacCer, GlcCer(d18:1/24:1), LacCer(d18:1/22:0) and Cer(d18:1/18:0).

In another preferred embodiment, the one or more lipid(s) whose increase in concentration is (are) compared to the control is (are) selected from (Table 8):

Total PC, PC 16:0/20:4, Cer(d18:0/24:0), Total LPC, CE 14:0, CE 20:3, CE 17:1, PC 16:0/20:3, LPC 18:1, PC 18:0/20:3, PC 18:0/18:1 and Cer(d18:0/22:0).

In a particularly preferred embodiment, the one or more lipid(s) whose decrease in concentration is (are) compared to the control is (are) selected from (Table 11):

Cer(d18:1/20:0), LacCer(d18:1/20:0), Cer(d18:1/24:1), LacCer(d18:1/24:1), LacCer(d18:1/22:0) and Cer(d18:1/18:0);

and the one or more lipid(s) whose increase in concentration is (are) compared to the control is (are) selected from (Table 11): PC 16:0/20:4 and Cer(d18:0/24:0).

In another alternative embodiment the invention relates to a method for evaluating the effectiveness of a treatment of CVD and/or one or more of its complications, such as AMI or CVD death, in a subject, comprising determining in a sample from said subject one or more lipid-lipid ratio(s), wherein (an) increased or decreased lipid-lipid ratio(s) in said sample, when compared to a control sample, is (are) indicative of effectiveness of said treatment, wherein the one or more lipid-lipid ratio(s) whose decrease is (are) compared to the control is (are) selected from (Tables 5a and 8a):

CE 16:0/CE 18:3, CE 18:2/CE 18:3, CE 19:1/Cer(d18:0/22:0), Cer(d18:1/16:0)/LPC 18:1, Cer(d18:1/16:0)/Total PC, Cer(d18:1/18:0)/LPC 16:0, Cer(d18:1/18:0)/LPC 18:1, Cer(d18:1/18:0)/PC 16:0/18:1, Cer(d18:1/18:0)/PC 16:0/20:3, Cer(d18:1/18:0)/PC 16:0/20:4, Cer(d18:1/18:0)/PC 18:0/20:4, Cer(d18:1/18:0)/Total LPC, Cer(d18:1/18:0)/Total PC, Cer(d18:1/20:0)/PC 16:0/18:1, Cer(d18:1/20:0)/PC 16:0/20:3, Cer(d18:1/20:0)/PC 16:0/20:4, Cer(d18:1/20:0)/PC 18:0/20:4, Cer(d18:1/20:0)/PC 18:1/18:1, Cer(d18:1/20:0)/Total PC, Cer(d18:1/22:0)/LPC 18:2, Cer(d18:1/22:0)/PC 16:0/20:3, Cer(d18:1/22:0)/PC 16:0/20:4, Cer(d18:1/22:0)/Total PC, Cer(d18:1/24:1)/LPC 18:1, Cer(d18:1/24:1)/LPC 18:2, Cer(d18:1/24:1)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 16:0/20:4, Cer(d18:1/24:1)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/20:3, Cer(d18:1/24:1)/PC 18:0/20:4, Cer(d18:1/24:1)/SM (d18:1/24:0) (d18:1/23:1-OH), Cer(d18:1/24:1)/Total CE, Cer(d18:1/24:1)/Total LPC, Cer(d18:1/24:1)/Total PC, Cer(d18:1/26:0)/PC O-40:0, GlcCer(d18:1/20:0)/PC 16:0/20:4, GlcCer(d18:1/20:0)/Total PC, GlcCer(d18:1/26:0)/Total CE, LacCer(d18:1/16:0)/Total LPC, LacCer(d18:1/18:0)/PC 16:0/18:1, LacCer(d18:1/18:0)/PC 16:0/20:3, LacCer(d18:1/18:0)/PC 18:0/18:1, LacCer(d18:1/18:0)/PC 18:0/20:3, LacCer(d18:1/18:0)/PC 18:1/18:1, LacCer(d18:1/18:0)/PC 18:1/18:2, LacCer(d18:1/18:0)/Total LPC, LacCer(d18:1/18:0)/Total PC, LacCer(d18:1/20:0)/PC 16:0/18:1, LacCer(d18:1/20:0)/PC 16:0/20:3, LacCer(d18:1/20:0)/PC 18:0/18:1, LacCer(d18:1/20:0)/PC 18:0/20:3 LacCer(d18:1/20:0)/PC 18:1/18:1, LacCer(d18:1/20:0)/PC 18:1/18:2, LacCer(d18:1/20:0)/PC 18:2/18:2, LacCer(d18:1/20:0)/SM (d18:1/17:1-OH), LacCer(d18:1/20:0)/SM (d18:1/18:0), LacCer(d18:1/20:0)/Total LPC, LacCer(d18:1/20:0)/Total PC, LacCer(d18:1/20:0)/Total SM, LacCer(d18:1/22:0)/PC 16:0/20:3, LacCer(d18:1/22:0)/PC 16:0/20:4, LacCer(d18:1/22:0)/PC 18:0/20:3, LacCer(d18:1/22:0)/SM (d18:1/14:0) (d18:1/13:1-OH), LacCer(d18:1/22:0)/Total LPC, LacCer(d18:1/22:0)/Total PC, LacCer(d18:1/24:0)/PC 16:0/20:3, LacCer(d18:1/24:0)/Total LPC, LacCer(d18:1/24:1)/Total LPC, LacCer(d18:1/24:1)/Total PC, LacCer(d18:1/24:1)/Total PC O, PC 16:0/16:0/PC 16:0/20:4, PC 16:0/16:0/Total PC, PC 16:0/18:2/Total PC, PC O-18:0/18:2-alkyl/PC O-36:5, PC O-32:0 (KDdiA-PC)/PC O-38:5, PS O-16:0/18:2-alkenyl/Total PS O, PS O-16:1/18:2-alkyl/Total PS O, PS O-18:0/18:2-alkenyl (PS O-18:1/18:2-alkyl)/Total PS O, PS O-18:2/16:0-alkenyl/Total PS O, Total Cer/Total PC, Total LacCer/Total PC and Total LacCer/Total PC O;

and wherein one or more lipid-lipid ratio(s) whose increase is (are) compared to the control is (are) selected from (Tables 5a and 8a):

CE 14:0/Cer(d18:1/24:1), CE 16:0/Cer(d18:1/24:1), CE 16:1/CE 19:1, CE 16:1/Cer(d18:1/18:0), CE 16:1/Cer(d18:1/20:0), CE 16:1/Cer(d18:1/24:1), CE 16:1/GlcCer(d18:1/18:0), CE 16:1/GlcCer(d18:1/20:0), CE 16:1/LacCer(d18:1/16:0), CE 16:1/LacCer(d18:1/18:0), CE 16:1/LacCer(d18:1/20:0), CE 16:1/LacCer(d18:1/22:0), CE 16:1/LacCer(d18:1/24:0), CE 16:1/PC 16:0/16:0, CE 16:1/Total LacCer, CE 17:1/Cer(d18:1/24:1), CE 17:1/GlcCer(d18:1/24:1), CE 17:1/LacCer(d18:1/18:0), CE 18:1/Total LacCer, CE 18:3/Cer(d18:1/16:0), CE 18:3/Cer(d18:1/18:0), CE 18:3/Cer(d18:1/20:0), CE 18:3/Cer(d18:1/22:0), CE 18:3/Cer(d18:1/24:0), CE 18:3/Cer(d18:1/24:1), CE 18:3/GlcCer(d18:1/18:0), CE 18:3/GlcCer(d18:1/20:0), CE 18:3/LacCer(d18:1/18:0), CE 18:3/LacCer(d18:1/20:0), CE 18:3/LacCer(d18:1/22:0), CE 18:3/LacCer(d18:1/24:0), CE 18:3/PC 16:0/16:0, CE 18:3/PC O-34:1, CE 18:3/PS O-16:0/18:1-alkenyl (PS O-16:1/18:1-alkyl), CE 18:3/PS O-16:0/18:2-alkenyl, CE 18:3/PS O-16:1/18:2-alkyl, CE 18:3/Total CE, CE 18:3/Total Cer, CE 18:3/Total LacCer, CE 20:3/Cer(d18:1/24:1), CE 20:3/LacCer(d18:1/20:0), Cer(d18:0/22:0)/Cer(d18:1/18:0), Cer(d18:0/22:0)/Cer(d18:1/20:0), Cer(d18:0/22:0)/Cer(d18:1/22:0), Cer(d18:0/22:0)/Cer(d18:1/24:1), Cer(d18:0/22:0)/LacCer(d18:1/24:0), Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/22:0)/Total CE, Cer(d18:0/24:0)/Cer(d18:1/16:0), Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/Cer(d18:1/20:0), Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/GlcCer(d18:1/20:0), Cer(d18:0/24:0)/LacCer(d18:1/24:0), Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/24:0)/SM (d18:1/17:0) (d18:1/16:1-OH), Cer(d18:0/24:0)/Total CE, Cer(d18:0/24:0)/Total Cer, Cer(d18:0/24:1)/Total CE, GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), GlcCer(d18:1/26:0)/LacCer(d18:1/22:0), LPC 16:0/LacCer(d18:1/20:0), LPC 16:0/LacCer(d18:1/22:0), LPC 16:0/LacCer(d18:1/24:1), LPC 16:0/Total LacCer, LPC 18:1/LacCer(d18:1/20:0), LPC 18:2/LacCer(d18:1/20:0), LPC 18:2/PS O-16:0/18:2-alkenyl, LPC 18:2/PS O-16:1/18:2-alkyl, PC 16:0/20:3/PS O-16:0/18:2-alkenyl, PC 16:0/20:3/PS O-16:1/18:2-alkyl, PC 18:1/18:2/PS O-16:0/18:2-alkenyl, PC 18:1/18:2/PS O-16:1/18:2-alkyl, SM (d18:1/24:0) (d18:1/23:1-OH)/Total Cer and Total LPC/Total LacCer.

In another alternative embodiment the invention relates to a method for evaluating the effectiveness of a treatment of CVD and/or one or more of its complications, such as AMI or CVD death, in a subject, wherein the subject is not undergoing statin treatment and wherein said method comprises comprising determining in a sample from said subject one or more lipid-lipid ratio(s), wherein (an) increased or decreased lipid-lipid ratio(s) in said sample, when compared to a control sample, is (are) indicative of effectiveness of said treatment, wherein the one or more lipid-lipid ratio(s) whose decrease is (are) compared to the control is (are) selected from (Tables 5b and 8b):

CE 16:0/CE 18:3, CE 18:0/CE 18:3, CE 18:2/CE 18:3, Cer(d18:1/16:0)/LPC 18:1, Cer(d18:1/16:0)/Total PC, Cer(d18:1/18:0)/LPC 18:1, Cer(d18:1/18:0)/PC 16:0/18:1, Cer(d18:1/18:0)/PC 16:0/20:3, Cer(d18:1/18:0)/PC 16:0/20:4, Cer(d18:1/18:0)/PC 18:0/18:1, Cer(d18:1/18:0)/PC 18:0/20:4, Cer(d18:1/18:0)/PC 18:1/18:1, Cer(d18:1/18:0)/SM (d18:1/14:0) (d18:1/13:1-OH), Cer(d18:1/18:0)/SM (d18:1/17:2-OH), Cer(d18:1/18:0)/SM (d18:1/18:1), Cer(d18:1/18:0)/Total CE, Cer(d18:1/18:0)/Total LPC, Cer(d18:1/18:0)/Total PC, Cer(d18:1/20:0)/PC 16:0/18:1, Cer(d18:1/20:0)/PC 16:0/20:3, Cer(d18:1/20:0)/PC 16:0/20:4, Cer(d18:1/20:0)/PC 18:0/18:1, Cer(d18:1/20:0)/PC 18:0/20:4, Cer(d18:1/20:0)/PC 18:1/18:1, Cer(d18:1/20:0)/SM (d18:1/14:0) (d18:1/13:1-OH), Cer(d18:1/20:0)/Total LPC, Cer(d18:1/20:0)/Total PC, Cer(d18:1/20:0)/Total PC O, Cer(d18:1/22:0)/LPC 18:2, Cer(d18:1/22:0)/PC 16:0/20:3, Cer(d18:1/22:0)/PC 16:0/20:4, Cer(d18:1/22:0)/PC 18:0/20:4, Cer(d18:1/22:0)/Total PC, Cer(d18:1/24:1)/LPC 18:1, Cer(d18:1/24:1)/LPC 18:2, Cer(d18:1/24:1)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:2, Cer(d18:1/24:1)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 16:0/20:4, Cer(d18:1/24:1)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:2, Cer(d18:1/24:1)/PC 18:0/20:3, Cer(d18:1/24:1)/PC 18:0/20:4, Cer(d18:1/24:1)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:2, Cer(d18:1/24:1)/PC O-40:3, Cer(d18:1/24:1)/SM (d18:1/17:1-OH), Cer(d18:1/24:1)/SM (d18:1/18:0), Cer(d18:1/24:1)/SM (d18:1/23:0) (d18:1/22:1-OH), Cer(d18:1/24:1)/SM (d18:1/24:0) (d18:1/23:1-OH), Cer(d18:1/24:1)/Total CE, Cer(d18:1/24:1)/Total LPC, Cer(d18:1/24:1)/Total PC, GlcCer(d18:1/26:0)/Total CE, GlcCer(d18:1/26:1)/Total CE, LacCer(d18:1/18:0)/Total PC, LacCer(d18:1/20:0)/PC 16:0/20:3, LacCer(d18:1/20:0)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 18:0/18:1, LacCer(d18:1/20:0)/PC 18:0/20:3, LacCer(d18:1/20:0)/PC 18:0/20:4, LacCer(d18:1/20:0)/PC 18:1/18:2, LacCer(d18:1/20:0)/SM (d18:1/17:1-OH), LacCer(d18:1/20:0)/SM (d18:1/18:0), LacCer(d18:1/20:0)/Total CE, LacCer(d18:1/20:0)/Total LPC, LacCer(d18:1/20:0)/Total PC, LacCer(d18:1/20:0)/Total SM, LacCer(d18:1/24:0)/Total LPC, PC 16:0/16:0/PC 16:0/20:4, PC 16:0/16:0/Total PC, PC O-32:0 (KDdiA-PC)/Total PC O, PS O-16:0/18:2-alkenyl/Total PC, PS O-16:0/18:2-alkenyl/Total PC O, PS O-16:0/18:2-alkenyl/Total PS O, PS O-16:1/18:2-alkyl/Total PC, PS O-16:1/18:2-alkyl/Total PC O, PS O-16:1/18:2-alkyl/Total PS O, PS O-18:2/16:0-alkenyl/Total PC O, PS O-18:2/16:0-alkenyl/Total PS O, SM (d18:1/17:0) (d18:1/16:1-OH)/Total PC O, Total Cer/Total PC, Total DAG/Total LPC, Total DAG/Total PC, Total DAG/Total PC O and Total LacCer/Total PC;

and wherein the one or more lipid-lipid ratio(s) whose increase(s) is (are) compared to the control is (are) selected from (Tables 5b and 8b):

CE 14:0/Cer(d18:1/18:0), CE 14:0/Cer(d18:1/24:1), CE 14:0/Total DAG, CE 15:0/Cer(d18:1/20:0), CE 16:0/Cer(d18:1/18:0), CE 16:0/Cer(d18:1/24:1), CE 16:1/CE 19:1, CE 16:1/Cer(d18:1/18:0), CE 16:1/Cer(d18:1/20:0), CE 16:1/Cer(d18:1/24:1), CE 16:1/GlcCer(d18:1/24:1), CE 16:1/LacCer(d18:1/18:0), CE 16:1/LacCer(d18:1/24:0), CE 16:1/Total LacCer, CE 17:1/Cer(d18:1/18:0), CE 17:1/Cer(d18:1/24:1), CE 18:2/Cer(d18:1/20:0), CE 18:2/Cer(d18:1/24:1), CE 18:3/Cer(d18:1/16:0), CE 18:3/Cer(d18:1/18:0), CE 18:3/Cer(d18:1/20:0), CE 18:3/Cer(d18:1/22:0), CE 18:3/Cer(d18:1/24:0), CE 18:3/Cer(d18:1/24:1), CE 18:3/GlcCer(d18:1/20:0), CE 18:3/LacCer(d18:1/20:0), CE 18:3/LacCer(d18:1/22:0), CE 18:3/LacCer(d18:1/24:0), CE 18:3/PC 16:0/16:0, CE 18:3/PC O-34:1, CE 18:3/PS O-16:0/18:2-alkenyl, CE 18:3/PS O-16:1/18:2-alkyl, CE 18:3/Total CE, CE 18:3/Total Cer, CE 18:3/Total DAG, CE 18:3/Total LacCer, CE 20:3/Cer(d18:1/24:1), CE 20:3/LacCer(d18:1/20:0), CE 20:4/Cer(d18:1/18:0), CE 20:4/Cer(d18:1/24:1), CE 20:4/GlcCer(d18:1/20:0), CE 20:4/GlcCer(d18:1/24:1), CE 20:4/LacCer(d18:1/20:0), CE 20:5/LacCer(d18:1/20:0), Cer(d18:0/22:0)/Cer(d18:1/18:0), Cer(d18:0/22:0)/Cer(d18:1/20:0), Cer(d18:0/22:0)/Cer(d18:1/24:1), Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/22:0)/Total CE, Cer(d18:0/22:0)/Total DAG, Cer(d18:0/22:0)/Total GlcCer, Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/Cer(d18:1/20:0), Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/P S O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/24:0)/Total CE, Cer(d18:0/24:0)/Total Cer, DAG 16:0/18:1/Total DAG, GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), LPC 16:0/LacCer(d18:1/24:0), LPC 18:1/LacCer(d18:1/20:0), LPC 18:2/LacCer(d18:1/20:0), PC 16:0/20:3/PS O-16:0/18:2-alkenyl, PC 16:0/20:3/PS O-16:1/18:2-alkyl, PC 16:0/20:4/PS O-16:0/18:2-alkenyl, PC 16:0/20:4/PS O-16:1/18:2-alkyl, PC 16:0/20:4/Total DAG, PC 18:0/20:3/PS O-16:0/18:2-alkenyl, PC 18:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-18:2/16:0-alkenyl, PC 18:0/20:4/PS O-16:0/18:2-alkenyl, PC 18:0/20:4/PS O-16:1/18:2-alkyl, PC 18:1/18:2/PS O-16:0/18:2-alkenyl, PC 18:1/18:2/PS O-16:1/18:2-alkyl, PC 18:1/18:2/Total Cer, PC O-40:3/PS O-18:2/16:0-alkenyl, SM (d18:1/23:0) (d18:1/22:1-OH)/Total DAG, SM (d18:1/24:0) (d18:1/23:1-OH)/Total Cer, Total CE/Total DAG and Total LPC/Total LacCer.

In a preferred embodiment (for subjects that are either undergoing or not undergoing statin treatment), the one or more lipid-lipid ratio(s) whose decrease is (are) compared to the control is (are) selected from (Table 9):

GlcCer(d18:1/26:1)/Total CE, Cer(d18:1/24:1)/Total PC, Cer(d18:1/24:1)/PC 16:0/20:4, Cer(d18:1/20:0)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 16:0/20:3, Total Cer/Total PC, Total LacCer/Total PC, LacCer(d18:1/20:0)/PC 18:1/18:2, PS O-16:0/18:2-alkenyl/Total PS O, Cer(d18:1/18:0)/PC 16:0/20:4, LacCer(d18:1/20:0)/Total LPC and LacCer(d18:1/20:0)/PC 16:0/20:4.

In another preferred embodiment, the one or more lipid-lipid ratio(s) whose increase is (are) compared to the control is (are) selected from (Table 9):

Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/22:0)/Cer(d18:1/24:1), DAG 16:0/18:1/Total DAG, Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/24:0)/Total CE, Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/Total Cer, Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/LacCer(d18:1/24:0), Cer(d18:0/22:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/22:0)/Cer(d18:1/20:0), Cer(d18:0/22:0)/P S O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl, GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), Total LPC/Total LacCer and GlcCer(d18:1/26:0)/LacCer(d18:1/22:0).

In a particularly preferred embodiment, the one or more lipid-lipid ratio(s) whose decrease is (are) compared to the control is (are) selected from (Table 11):

Cer(d18:1/24:1)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 16:0/20:3, PS O-16:0/18:2-alkenyl/Total PS O and Cer(d18:1/18:0)/PC 16:0/20:4;

and the one or more lipid-lipid ratio(s) whose increase is (are) compared to the control is (are) selected from (Table 11):

GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), DAG 16:0/18:1/Total DAG, Cer(d18:0/24:0)/Total Cer, Total LPC/Total LacCer, GlcCer(d18:1/26:0)/LacCer(d18:1/22:0), Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl and Cer(d18:0/24:0)/LacCer(d18:1/24:0).

In yet another alternative embodiment the invention relates to a method for evaluating the effectiveness of a treatment of CVD and/or one or more of its complications, such as AMI or CVD death, in a subject, comprising determining in a sample from said subject one or more lipid-clinical concentration ratio(s), wherein (an) increased or decreased lipid-clinical concentration ratio(s) in said sample, when compared to a control sample, is (are) indicative of effectiveness of said treatment, wherein the one or more lipid-clinical concentration ratio(s) whose decrease(s) is (are) compared to the control is (are) selected from (Tables 6a and 9a):

Cer(d18:1/16:0)/triglycerides, Cer(d18:1/18:0)/apolipoprotein A-I, Cer(d18:1/18:0)/apolipoprotein B, Cer(d18:1/18:0)/HDL cholesterol, Cer(d18:1/18:0)/total cholesterol, Cer(d18:1/18:0)/total-c/HDL-c, Cer(d18:1/18:0)/triglycerides, Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/20:0)/apolipoprotein B, Cer(d18:1/20:0)/HDL cholesterol, Cer(d18:1/20:0)/total cholesterol, Cer(d18:1/20:0)/triglycerides, Cer(d18:1/22:0)/apolipoprotein B, Cer(d18:1/22:0)/LDL cholesterol, Cer(d18:1/22:0)/triglycerides, Cer(d18:1/24:1)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein B, Cer(d18:1/24:1)/LDL cholesterol, Cer(d18:1/24:1)/total cholesterol, Cer(d18:1/24:1)/total-c/HDL-c, Cer(d18:1/24:1)/triglycerides, GlcCer(d18:1/24:0)/total cholesterol, LacCer(d18:1/16:0)/triglycerides, LacCer(d18:1/18:0)/apolipoprotein A-I, LacCer(d18:1/18:0)/apolipoprotein B, LacCer(d18:1/18:0)/HDL cholesterol, LacCer(d18:1/18:0)/LDL cholesterol, LacCer(d18:1/18:0)/total cholesterol, LacCer(d18:1/18:0)/total-c/HDL-c, LacCer(d18:1/18:0)/triglycerides, LacCer(d18:1/20:0)/apoA1/apoB, LacCer(d18:1/20:0)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein B, LacCer(d18:1/20:0)/HDL cholesterol, LacCer(d18:1/20:0)/LDL cholesterol, LacCer(d18:1/20:0)/LDL-c/HDL-c, LacCer(d18:1/20:0)/total cholesterol, LacCer(d18:1/20:0)/total-c/HDL-c, LacCer(d18:1/20:0)/triglycerides, LacCer(d18:1/22:0)/apoA/apoB, LacCer(d18:1/22:0)/apolipoprotein A-I, LacCer(d18:1/22:0)/apolipoprotein B, LacCer(d18:1/22:0)/HDL cholesterol, LacCer(d18:1/22:0)/LDL cholesterol, LacCer(d18:1/22:0)/total cholesterol, LacCer(d18:1/22:0)/total-c/HDL-c, LacCer(d18:1/22:0)/triglycerides, LacCer(d18:1/24:0)/apoA/apoB, LacCer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/24:1)/apolipoprotein B, LacCer(d18:1/24:1)/total cholesterol, LacCer(d18:1/24:1)/triglycerides, PC O-32:0 (KDdiA-PC)/apolipoprotein A-I, PC O-32:0 (KDdiA-PC)/triglycerides, PC O-34:1/triglycerides, PS O-16:0/18:1-alkenyl (PS O-16:1/18:1-alkyl)/triglycerides, PS O-16:0/18:2-alkenyl/triglycerides, PS O-16:1/18:2-alkyl/triglycerides, PS O-18:2/16:0-alkenyl/HDL cholesterol, PS O-18:2/16:0-alkenyl/triglycerides, Total Cer/apolipoprotein A-I, Total Cer/total cholesterol, Total Cer/triglycerides, Total GlcCer/apolipoprotein B, Total GlcCer/total cholesterol, Total LacCer/apolipoprotein A-I, Total LacCer/apolipoprotein B, Total LacCer/total cholesterol and Total LacCer/triglycerides;

and wherein the one or more lipid-clinical concentration ratio(s) whose increase is (are) compared to the control is (are) selected from Tables 6a and 9a:

CE 14:0/apolipoprotein B, CE 14:0/LDL cholesterol, CE 14:0/LDL-c/HDL-c, CE 14:0/total cholesterol, CE 14:0/total-c/HDL-c, CE 16:1/apolipoprotein B, CE 16:1/HDL cholesterol, CE 16:1/LDL cholesterol, CE 16:1/total cholesterol, CE 17:1/LDL-c/HDL-c, CE 18:3/apoA1/apoB, CE 18:3/apolipoprotein A-I, CE 18:3/apolipoprotein B, CE 18:3/HDL cholesterol, CE 18:3/LDL cholesterol, CE 18:3/LDL-c/HDL-c, CE 18:3/total cholesterol, CE 18:3/total-c/HDL-c, CE 20:3/apolipoprotein B, CE 20:3/LDL-c/HDL-c, CE 20:3/total-c/HDL-c, CE 20:5/apolipoprotein B, CE 20:5/HDL cholesterol, CE 20:5/LDL cholesterol, Cer(d18:0/22:0)/apolipoprotein B, Cer(d18:0/22:0)/LDL-c/HDL-c, Cer(d18:0/22:0)/total-c/HDL-c, Cer(d18:0/24:0)/apolipoprotein A-I, Cer(d18:0/24:0)/apolipoprotein B, Cer(d18:0/24:0)/HDL cholesterol, Cer(d18:0/24:0)/LDL cholesterol, Cer(d18:0/24:0)/LDL-c/HDL-c, Cer(d18:0/24:0)/total cholesterol, Cer(d18:0/24:0)/total-c/HDL-c, LPC 18:2/apoA1/apoB, LPC 18:2/apolipoprotein B, LPC 18:2/HDL cholesterol, LPC 18:2/LDL cholesterol, LPC 18:2/LDL-c/HDL-c, LPC 18:2/total cholesterol, PC 16:0/20:3/apolipoprotein B, PC 16:0/20:3/HDL cholesterol, PC 16:0/20:3/LDL-c/HDL-c, PC 16:0/20:3/total-c/HDL-c, PC 16:0/20:4/apolipoprotein A-I, PC 16:0/20:4/apolipoprotein B, PC 16:0/20:4/LDL cholesterol, PC 16:0/20:4/LDL-c/HDL-c, PC 16:0/20:4/total cholesterol, PC 16:0/20:4/total-c/HDL-c, PC 18:0/18:1/LDL-c/HDL-c, PC 18:0/20:3/LDL-c/HDL-c, PC 18:0/20:3/total-c/HDL-c, PC 18:0/20:4/apoA1/apoB, Total LPC/LDL-c/HDL-c, Total LPC/total-c/HDL-c, Total PC/apolipoprotein B, Total PC/LDL-c/HDL-c, Total PC/total cholesterol and Total PC/total-c/HDL-c.

In yet another alternative embodiment the invention relates to a method for evaluating the effectiveness of a treatment of CVD and/or one or more of its complications, such as AMI or CVD death, in a subject, wherein the subject is not undergoing statin treatment and wherein said method comprises determining in a sample from said subject one or more lipid-clinical concentration ratio(s), wherein (an) increased or decreased lipid-clinical concentration ratio(s) in said sample, when compared to a control sample, is (are) indicative of effectiveness of said treatment, wherein the one or more lipid-clinical concentration ratio(s) whose decrease(s) is (are) compared to the control is (are) selected from (Tables 6b and 9b):

Cer(d18:1/16:0)/apolipoprotein A-I, Cer(d18:1/16:0)/LDL cholesterol, Cer(d18:1/16:0)/triglycerides, Cer(d18:1/18:0)/apolipoprotein A-I, Cer(d18:1/18:0)/apolipoprotein B, Cer(d18:1/18:0)/HDL cholesterol, Cer(d18:1/18:0)/total cholesterol, Cer(d18:1/18:0)/total-c/HDL-c, Cer(d18:1/18:0)/triglycerides, Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/20:0)/apolipoprotein B, Cer(d18:1/20:0)/HDL cholesterol, Cer(d18:1/20:0)/LDL cholesterol, Cer(d18:1/20:0)/total cholesterol, Cer(d18:1/20:0)/total-c/HDL-c, Cer(d18:1/20:0)/triglycerides, Cer(d18:1/22:0)/apolipoprotein A-I, Cer(d18:1/22:0)/apolipoprotein B, Cer(d18:1/22:0)/LDL cholesterol, Cer(d18:1/22:0)/total cholesterol, Cer(d18:1/22:0)/triglycerides, Cer(d18:1/24:0)/apolipoprotein A-I, Cer(d18:1/24:0)/apolipoprotein B, Cer(d18:1/24:0)/LDL cholesterol, Cer(d18:1/24:0)/total cholesterol, Cer(d18:1/24:1)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein B, Cer(d18:1/24:1)/HDL cholesterol, Cer(d18:1/24:1)/LDL cholesterol, Cer(d18:1/24:1)/LDL-c/HDL-c, Cer(d18:1/24:1)/total cholesterol, Cer(d18:1/24:1)/total-c/HDL-c, Cer(d18:1/24:1)/triglycerides, GlcCer(d18:1/20:0)/apolipoprotein B, GlcCer(d18:1/20:0)/total cholesterol, GlcCer(d18:1/24:1)/apolipoprotein B, GlcCer(d18:1/26:1)/apolipoprotein A-I, LacCer(d18:1/16:0)/triglycerides, LacCer(d18:1/18:0)/apolipoprotein A-I, LacCer(d18:1/18:0)/apolipoprotein B, LacCer(d18:1/18:0)/total cholesterol, LacCer(d18:1/20:0)/apoA1/apoB, LacCer(d18:1/20:0)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein B, LacCer(d18:1/20:0)/HDL cholesterol, LacCer(d18:1/20:0)/LDL cholesterol, LacCer(d18:1/20:0)/LDL-c/HDL-c, LacCer(d18:1/20:0)/total cholesterol, LacCer(d18:1/20:0)/total-c/HDL-c, LacCer(d18:1/20:0)/triglycerides, LacCer(d18:1/22:0)/apoA1/apoB, LacCer(d18:1/22:0)/apolipoprotein A-I, LacCer(d18:1/22:0)/apolipoprotein B, LacCer(d18:1/22:0)/HDL cholesterol, LacCer(d18:1/22:0)/LDL cholesterol, LacCer(d18:1/22:0)/total cholesterol, LacCer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/24:1)/apolipoprotein B, LacCer(d18:1/24:1)/total cholesterol, LacCer(d18:1/24:1)/triglycerides, PC O-34:1/apolipoprotein B, PS O-16:0/18:2-alkenyl/triglycerides, PS O-16:1/18:2-alkyl/triglycerides, Total Cer/apolipoprotein A-I, Total Cer/apolipoprotein B, Total Cer/LDL cholesterol, Total Cer/total cholesterol, Total DAG/apolipoprotein A-I, Total DAG/triglycerides, Total GlcCer/apolipoprotein B, Total LacCer/apolipoprotein A-I, Total LacCer/apolipoprotein B and Total LacCer/total cholesterol.

and wherein the one or more lipid-clinical concentration ratio(s) whose increase is (are) compared to the control is (are) selected from (Tables 6b and 9b):

CE 14:0/apoA1/apoB, CE 14:0/apolipoprotein B, CE 14:0/LDL-c/HDL-c, CE 14:0/total-c/HDL-c, CE 16:1/apoA1/apoB, CE 18:3/apoA1/apoB, CE 18:3/apolipoprotein A-I, CE 18:3/apolipoprotein B, CE 18:3/HDL cholesterol, CE 18:3/LDL cholesterol, CE 18:3/LDL-c/HDL-c, CE 18:3/total cholesterol, CE 18:3/total-c/HDL-c, CE 20:5/triglycerides, Cer(d18:0/24:0)/apolipoprotein B, Cer(d18:0/24:0)/total cholesterol, Cer(d18:0/24:0)/total-c/HDL-c, PC 18:0/20:4/apoA1/apoB, PC O-38:6/apolipoprotein A-I, Total LPC/apoA1/apoB and Total PC/apolipoprotein A-I.

In a preferred embodiment (for subjects that are either undergoing or not undergoing statin treatment), the one or more lipid-clinical concentration ratio(s) whose decrease is (are) compared to the control is (are) selected from (Table 10):

Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein A-I, Total Cer/apolipoprotein A-I, Total LacCer/apolipoprotein A-I, Cer(d18:1/18:0)/apolipoprotein A-I, LacCer(d18:1/22:0)/apolipoprotein A-I, LacCer(d18:1/20:0)/HDL cholesterol and Cer(d18:1/24:1)/apolipoprotein B.

In another preferred embodiment, the one or more lipid-clinical concentration ratio(s) whose increase is (are) compared to the control is (are) selected from (Table 10):

Cer(d18:0/24:0)/apolipoprotein B, Cer(d18:0/24:0)/total cholesterol, Cer(d18:0/24:0)/apolipoprotein B, PC 16:0/20:4/apolipoprotein B and Cer(d18:0/24:0)/apolipoprotein A-I.

In a particularly preferred embodiment, the lipid-clinical concentration ratio(s) whose decrease(s) is (are) compared to the control are selected from (Table 11):

Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein A-I, Total LacCer/apolipoprotein A-I, LacCer(d18:1/20:0)/HDL cholesterol and Cer(d18:1/18:0)/apolipoprotein A-I.

For the purposes of the invention, and particularly for lipid-clinical concentration ratios, an Apolipoprotein A-I measurement may alternatively be an Apolipoprotein A-II measurement.

In yet another aspect the invention relates to a method of choosing an appropriate treatment of CVD and/or one or more of its complications, such as AMI or CVD death, in a subject, comprising determining in a sample from said subject the concentration of one or more lipid(s), wherein (an) increased or decreased concentration(s) in said sample, when compared to a control sample, is (are) indicative of said subject being in need of treatment or a change in, or supplementation of, an already administered treatment, wherein the one or more lipid(s) whose increase(s) in concentration is (are) compared to the control is (are) selected from (Tables 4a and 7a):

Cer(d18:1/18:0), Cer(d18:1/20:0), Cer(d18:1/24:1), GlcCer(d18:1/18:0), LacCer(d18:1/18:0), LacCer(d18:1/20:0), LacCer(d18:1/22:0), LacCer(d18:1/24:0), PC O-32:0 (KDdiA-PC), PS O-16:0/18:2-alkenyl, PS O-16:1/18:2-alkyl, PS O-18:0/18:2-alkenyl (PS O-18:1/18:2-alkyl), PS O-18:2/16:0-alkenyl and Total LacCer;

and wherein the one or more lipid(s) whose decrease(s) in concentration is (are) compared to the control is (are) selected from (Tables 4a and 7a):

CE 14:0, CE 16:0, CE 17:1, CE 20:3, Cer(d18:0/22:0), Cer(d18:0/24:0), LPC 18:1, PC 16:0/18:2, PC 16:0/20:3, PC 16:0/20:4, PC 16:0/22:6, PC 18:0/18:1, PC 18:0/20:3, PC 18:0/20:4, PC 18:1/18:2, SM (d18:1/14:0) (d18:1/13:1-OH), SM (d18:1/23:0) (d18:1/22:1-OH), SM (d18:1/24:0) (d18:1/23:1-OH), Total CE, Total LPC and Total PC.

In yet another aspect the invention relates to a method of choosing an appropriate treatment of CVD and/or one or more of its complications, such as AMI or CVD death, in a subject, wherein the subject is not undergoing statin treatment and wherein said method comprises determining in a sample from said subject the concentration of one or more lipid(s), wherein (an) increased or decreased concentration(s) in said sample, when compared to a control sample, is (are) indicative of said subject being in need of treatment or a change in, or supplementation of, an already administered treatment, wherein the one or more lipid(s) whose increase(s) in concentration is (are) compared to the control is (are) selected from (Tables 4b and 7b):

Cer(d18:1/16:0), Cer(d18:1/18:0), Cer(d18:1/20:0), Cer(d18:1/22:0), Cer(d18:1/24:1), Cer(d18:1/26:1), GlcCer(d18:1/18:0), GlcCer(d18:1/20:0), GlcCer(d18:1/24:1), GlcCer(d18:1/26:1), LacCer(d18:1/18:0), LacCer(d18:1/20:0), LacCer(d18:1/22:0), LacCer(d18:1/24:0), LacCer(d18:1/24:1), PC O-32:0 (KDdiA-PC), PS O-16:0/18:2-alkenyl, PS O-16:1/18:2-alkyl, PS O-18:2/16:0-alkenyl, Total Cer, Total DAG and Total LacCer;

and wherein the one or more lipid(s) whose decrease(s) in concentration is (are) compared to the control is (are) selected from (Tables 4b and 7b):

CE 14:0, CE 17:1, CE 20:3, Cer(d18:0/24:0), LPC 18:1, PC 16:0/20:3, PC 16:0/20:4, PC 18:0/20:4, PC O-40:3, SM (d18:1/14:0) (d18:1/13:1-OH), Total LPC and Total PC.

In a preferred embodiment (for subjects that are either undergoing or not undergoing statin treatment), the one or more lipid(s) whose increase in concentration is (are) compared to the control is (are) selected from (Table 8):

Cer(d18:1/20:0), LacCer(d18:1/20:0), Cer(d18:1/24:1), LacCer(d18:1/24:1), PS O-18:2/16:0-alkenyl, PS O-16:1/18:2-alkyl, Total Cer, Total LacCer, GlcCer(d18:1/24:1), LacCer(d18:1/22:0) and Cer(d18:1/18:0).

In another preferred embodiment, the one or more lipid(s) whose decrease in concentration is (are) compared to the control is (are) selected from (Table 8):

Total PC, PC 16:0/20:4, Cer(d18:0/24:0), Total LPC, CE 14:0, CE 20:3, CE 17:1, PC 16:0/20:3, LPC 18:1, PC 18:0/20:3, PC 18:0/18:1 and Cer(d18:0/22:0).

In a particularly preferred embodiment, the one or more lipid(s) whose increase in concentration is (are) compared to the control is (are) selected from (Table 11):

Cer(d18:1/20:0), LacCer(d18:1/20:0), Cer(d18:1/24:1), LacCer(d18:1/24:1), LacCer(d18:1/22:0) and Cer(d18:1/18:0);

and the one or more lipid(s) whose decrease in concentration is (are) compared to the control is (are) selected from (Table 11): PC 16:0/20:4 and Cer(d18:0/24:0).

In an alternative embodiment the invention relates to a method of choosing an appropriate treatment of CVD and/or one or more of its complications, such as AMI or CVD death, in a subject, comprising determining in a sample from said subject one or more lipid-lipid ratio(s), wherein (an) increased or decreased lipid-lipid ratio(s) in said sample, when compared to a control sample, is (are) indicative of said subject being in need of treatment or a change in, or supplementation of, an already administered treatment, wherein the one or more lipid-lipid ratio(s) whose increase(s) is (are) compared to the control is (are) selected from (Tables 5a and 8a):

CE 16:0/CE 18:3, CE 18:2/CE 18:3, CE 19:1/Cer(d18:0/22:0), Cer(d18:1/16:0)/LPC 18:1, Cer(d18:1/16:0)/Total PC, Cer(d18:1/18:0)/LPC 16:0, Cer(d18:1/18:0)/LPC 18:1, Cer(d18:1/18:0)/PC 16:0/18:1, Cer(d18:1/18:0)/PC 16:0/20:3, Cer(d18:1/18:0)/PC 16:0/20:4, Cer(d18:1/18:0)/PC 18:0/20:4, Cer(d18:1/18:0)/Total LPC, Cer(d18:1/18:0)/Total PC, Cer(d18:1/20:0)/PC 16:0/18:1, Cer(d18:1/20:0)/PC 16:0/20:3, Cer(d18:1/20:0)/PC 16:0/20:4, Cer(d18:1/20:0)/PC 18:0/20:4, Cer(d18:1/20:0)/PC 18:1/18:1, Cer(d18:1/20:0)/Total PC, Cer(d18:1/22:0)/LPC 18:2, Cer(d18:1/22:0)/PC 16:0/20:3, Cer(d18:1/22:0)/PC 16:0/20:4, Cer(d18:1/22:0)/Total PC, Cer(d18:1/24:1)/LPC 18:1, Cer(d18:1/24:1)/LPC 18:2, Cer(d18:1/24:1)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 16:0/20:4, Cer(d18:1/24:1)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/20:3, Cer(d18:1/24:1)/PC 18:0/20:4, Cer(d18:1/24:1)/SM (d18:1/24:0) (d18:1/23:1-OH), Cer(d18:1/24:1)/Total CE, Cer(d18:1/24:1)/Total LPC, Cer(d18:1/24:1)/Total PC, Cer(d18:1/26:0)/PC O-40:0, GlcCer(d18:1/20:0)/PC 16:0/20:4, GlcCer(d18:1/20:0)/Total PC, GlcCer(d18:1/26:0)/Total CE, LacCer(d18:1/16:0)/Total LPC, LacCer(d18:1/18:0)/PC 16:0/18:1, LacCer(d18:1/18:0)/PC 16:0/20:3, LacCer(d18:1/18:0)/PC 18:0/18:1, LacCer(d18:1/18:0)/PC 18:0/20:3, LacCer(d18:1/18:0)/PC 18:1/18:1, LacCer(d18:1/18:0)/PC 18:1/18:2, LacCer(d18:1/18:0)/Total LPC, LacCer(d18:1/18:0)/Total PC, LacCer(d18:1/20:0)/PC 16:0/18:1, LacCer(d18:1/20:0)/PC 16:0/20:3, LacCer(d18:1/20:0)/PC 18:0/18:1, LacCer(d18:1/20:0)/PC 18:0/20:3 LacCer(d18:1/20:0)/PC 18:1/18:1, LacCer(d18:1/20:0)/PC 18:1/18:2, LacCer(d18:1/20:0)/PC 18:2/18:2, LacCer(d18:1/20:0)/SM (d18:1/17:1-OH), LacCer(d18:1/20:0)/SM (d18:1/18:0), LacCer(d18:1/20:0)/Total LPC, LacCer(d18:1/20:0)/Total PC, LacCer(d18:1/20:0)/Total SM, LacCer(d18:1/22:0)/PC 16:0/20:3, LacCer(d18:1/22:0)/PC 16:0/20:4, LacCer(d18:1/22:0)/PC 18:0/20:3, LacCer(d18:1/22:0)/SM (d18:1/14:0) (d18:1/13:1-OH), LacCer(d18:1/22:0)/Total LPC, LacCer(d18:1/22:0)/Total PC, LacCer(d18:1/24:0)/PC 16:0/20:3, LacCer(d18:1/24:0)/Total LPC, LacCer(d18:1/24:1)/Total LPC, LacCer(d18:1/24:1)/Total PC, LacCer(d18:1/24:1)/Total PC O, PC 16:0/16:0/PC 16:0/20:4, PC 16:0/16:0/Total PC, PC 16:0/18:2/Total PC, PC O-18:0/18:2-alkyl/PC O-36:5, PC O-32:0 (KDdiA-PC)/PC O-38:5, PS O-16:0/18:2-alkenyl/Total PS O, PS O-16:1/18:2-alkyl/Total PS O, PS O-18:0/18:2-alkenyl (PS O-18:1/18:2-alkyl)/Total PS O, PS O-18:2/16:0-alkenyl/Total PS O, Total Cer/Total PC, Total LacCer/Total PC and Total LacCer/Total PC O;

and wherein one or more lipid-lipid ratio(s) whose decrease is (are) compared to the control is (are) selected from (Tables 5a and 8a):

CE 14:0/Cer(d18:1/24:1), CE 16:0/Cer(d18:1/24:1), CE 16:1/CE 19:1, CE 16:1/Cer(d18:1/18:0), CE 16:1/Cer(d18:1/20:0), CE 16:1/Cer(d18:1/24:1), CE 16:1/GlcCer(d18:1/18:0), CE 16:1/GlcCer(d18:1/20:0), CE 16:1/LacCer(d18:1/16:0), CE 16:1/LacCer(d18:1/18:0), CE 16:1/LacCer(d18:1/20:0), CE 16:1/LacCer(d18:1/22:0), CE 16:1/LacCer(d18:1/24:0), CE 16:1/PC 16:0/16:0, CE 16:1/Total LacCer, CE 17:1/Cer(d18:1/24:1), CE 17:1/GlcCer(d18:1/24:1), CE 17:1/LacCer(d18:1/18:0), CE 18:1/Total LacCer, CE 18:3/Cer(d18:1/16:0), CE 18:3/Cer(d18:1/18:0), CE 18:3/Cer(d18:1/20:0), CE 18:3/Cer(d18:1/22:0), CE 18:3/Cer(d18:1/24:0), CE 18:3/Cer(d18:1/24:1), CE 18:3/GlcCer(d18:1/18:0), CE 18:3/GlcCer(d18:1/20:0), CE 18:3/LacCer(d18:1/18:0), CE 18:3/LacCer(d18:1/20:0), CE 18:3/LacCer(d18:1/22:0), CE 18:3/LacCer(d18:1/24:0), CE 18:3/PC 16:0/16:0, CE 18:3/PC O-34:1, CE 18:3/PS O-16:0/18:1-alkenyl (PS O-16:1/18:1-alkyl), CE 18:3/PS O-16:0/18:2-alkenyl, CE 18:3/PS O-16:1/18:2-alkyl, CE 18:3/Total CE, CE 18:3/Total Cer, CE 18:3/Total LacCer, CE 20:3/Cer(d18:1/24:1), CE 20:3/LacCer(d18:1/20:0), Cer(d18:0/22:0)/Cer(d18:1/18:0), Cer(d18:0/22:0)/Cer(d18:1/20:0), Cer(d18:0/22:0)/Cer(d18:1/22:0), Cer(d18:0/22:0)/Cer(d18:1/24:1), Cer(d18:0/22:0)/LacCer(d18:1/24:0), Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/22:0)/Total CE, Cer(d18:0/24:0)/Cer(d18:1/16:0), Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/Cer(d18:1/20:0), Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/GlcCer(d8:1/20:0), Cer(d18:0/24:0)/LacCer(d18:1/24:0), Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/24:0)/SM (d18:1/17:0) (d18:1/16:1-OH), Cer(d18:0/24:0)/Total CE, Cer(d18:0/24:0)/Total Cer, Cer(d18:0/24:1)/Total CE, GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), GlcCer(d18:1/26:0)/LacCer(d18:1/22:0), LPC 16:0/LacCer(d18:1/20:0), LPC 16:0/LacCer(d18:1/22:0), LPC 16:0/LacCer(d18:1/24:1), LPC 16:0/Total LacCer, LPC 18:1/LacCer(d18:1/20:0), LPC 18:2/LacCer(d18:1/20:0), LPC 18:2/PS O-16:0/18:2-alkenyl, LPC 18:2/PS O-16:1/18:2-alkyl, PC 16:0/20:3/PS O-16:0/18:2-alkenyl, PC 16:0/20:3/PS O-16:1/18:2-alkyl, PC 18:1/18:2/PS O-16:0/18:2-alkenyl, PC 18:1/18:2/PS O-16:1/18:2-alkyl, SM (d18:1/24:0) (d18:1/23:1-OH)/Total Cer and Total LPC/Total LacCer.

In an alternative embodiment the invention relates to a method of choosing an appropriate treatment of CVD and/or one or more of its complications, such as AMI or CVD death, in a subject, wherein the subject is not undergoing statin treatment and wherein said method comprises determining in a sample from said subject one or more lipid-lipid ratio(s), wherein (an) increased or decreased lipid-lipid ratio(s) in said sample, when compared to a control sample, is (are) indicative of said subject being in need of treatment or a change in, or supplementation of, an already administered treatment, wherein the one or more lipid-lipid ratio(s) whose increase(s) is (are) compared to the control is (are) selected from (Tables 5b and 8b):

CE 16:0/CE 18:3, CE 18:0/CE 18:3, CE 18:2/CE 18:3, Cer(d18:1/16:0)/LPC 18:1, Cer(d18:1/16:0)/Total PC, Cer(d18:1/18:0)/LPC 18:1, Cer(d18:1/18:0)/PC 16:0/18:1, Cer(d18:1/18:0)/PC 16:0/20:3, Cer(d18:1/18:0)/PC 16:0/20:4, Cer(d18:1/18:0)/PC 18:0/18:1, Cer(d18:1/18:0)/PC 18:0/20:4, Cer(d18:1/18:0)/PC 18:1/18:1, Cer(d18:1/18:0)/SM (d18:1/14:0) (d18:1/13:1-OH), Cer(d18:1/18:0)/SM (d18:1/17:2-OH), Cer(d18:1/18:0)/SM (d18:1/18:1), Cer(d18:1/18:0)/Total CE, Cer(d18:1/18:0)/Total LPC, Cer(d18:1/18:0)/Total PC, Cer(d18:1/20:0)/PC 16:0/18:1, Cer(d18:1/20:0)/PC 16:0/20:3, Cer(d18:1/20:0)/PC 16:0/20:4, Cer(d18:1/20:0)/PC 18:0/18:1, Cer(d18:1/20:0)/PC 18:0/20:4, Cer(d18:1/20:0)/PC 18:1/18:1, Cer(d18:1/20:0)/SM (d18:1/14:0) (d18:1/13:1-OH), Cer(d18:1/20:0)/Total LPC, Cer(d18:1/20:0)/Total PC, Cer(d18:1/20:0)/Total PC O, Cer(d18:1/22:0)/LPC 18:2, Cer(d18:1/22:0)/PC 16:0/20:3, Cer(d18:1/22:0)/PC 16:0/20:4, Cer(d18:1/22:0)/PC 18:0/20:4, Cer(d18:1/22:0)/Total PC, Cer(d18:1/24:1)/LPC 18:1, Cer(d18:1/24:1)/LPC 18:2, Cer(d18:1/24:1)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:2, Cer(d18:1/24:1)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 16:0/20:4, Cer(d18:1/24:1)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:2, Cer(d18:1/24:1)/PC 18:0/20:3, Cer(d18:1/24:1)/PC 18:0/20:4, Cer(d18:1/24:1)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:2, Cer(d18:1/24:1)/PC O-40:3, Cer(d18:1/24:1)/SM (d18:1/17:1-OH), Cer(d18:1/24:1)/SM (d18:1/18:0), Cer(d18:1/24:1)/SM (d18:1/23:0) (d18:1/22:1-OH), Cer(d18:1/24:1)/SM (d18:1/24:0) (d18:1/23:1-OH), Cer(d18:1/24:1)/Total CE, Cer(d18:1/24:1)/Total LPC, Cer(d18:1/24:1)/Total PC, GlcCer(d18:1/26:0)/Total CE, GlcCer(d18:1/26:1)/Total CE, LacCer(d18:1/18:0)/Total PC, LacCer(d18:1/20:0)/PC 16:0/20:3, LacCer(d18:1/20:0)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 18:0/18:1, LacCer(d18:1/20:0)/PC 18:0/20:3, LacCer(d18:1/20:0)/PC 18:0/20:4, LacCer(d18:1/20:0)/PC 18:1/18:2, LacCer(d18:1/20:0)/SM (d18:1/17:1-OH), LacCer(d18:1/20:0)/SM (d18:1/18:0), LacCer(d18:1/20:0)/Total CE, LacCer(d18:1/20:0)/Total LPC, LacCer(d18:1/20:0)/Total PC, LacCer(d18:1/20:0)/Total SM, LacCer(d18:1/24:0)/Total LPC, PC 16:0/16:0/PC 16:0/20:4, PC 16:0/16:0/Total PC, PC O-32:0 (KDdiA-PC)/Total PC O, PS O-16:0/18:2-alkenyl/Total PC, PS O-16:0/18:2-alkenyl/Total PC O, PS O-16:0/18:2-alkenyl/Total PS O, PS O-16:1/18:2-alkyl/Total PC, PS O-16:1/18:2-alkyl/Total PC O, PS O-16:1/18:2-alkyl/Total PS O, PS O-18:2/16:0-alkenyl/Total PC O, PS O-18:2/16:0-alkenyl/Total PS O, SM (d18:1/17:0) (d18:1/16:1-OH)/Total PC O, Total Cer/Total PC, Total DAG/Total LPC, Total DAG/Total PC, Total DAG/Total PC O and Total LacCer/Total PC;

and wherein the one or more lipid-lipid ratio(s) whose decrease(s) is (are) compared to the control is (are) selected from (Tables 5b and 8b):

CE 14:0/Cer(d18:1/18:0), CE 14:0/Cer(d18:1/24:1), CE 14:0/Total DAG, CE 15:0/Cer(d18:1/20:0), CE 16:0/Cer(d18:1/18:0), CE 16:0/Cer(d18:1/24:1), CE 16:1/CE 19:1, CE 16:1/Cer(d18:1/18:0), CE 16:1/Cer(d18:1/20:0), CE 16:1/Cer(d18:1/24:1), CE 16:1/GlcCer(d18:1/24:1), CE 16:1/LacCer(d18:1/18:0), CE 16:1/LacCer(d18:1/24:0), CE 16:1/Total LacCer, CE 17:1/Cer(d18:1/18:0), CE 17:1/Cer(d18:1/24:1), CE 18:2/Cer(d18:1/20:0), CE 18:2/Cer(d18:1/24:1), CE 18:3/Cer(d18:1/16:0), CE 18:3/Cer(d18:1/18:0), CE 18:3/Cer(d18:1/20:0), CE 18:3/Cer(d18:1/22:0), CE 18:3/Cer(d18:1/24:0), CE 18:3/Cer(d18:1/24:1), CE 18:3/GlcCer(d18:1/20:0), CE 18:3/LacCer(d18:1/20:0), CE 18:3/LacCer(d18:1/22:0), CE 18:3/LacCer(d18:1/24:0), CE 18:3/PC 16:0/16:0, CE 18:3/PC O-34:1, CE 18:3/PS O-16:0/18:2-alkenyl, CE 18:3/PS O-16:1/18:2-alkyl, CE 18:3/Total CE, CE 18:3/Total Cer, CE 18:3/Total DAG, CE 18:3/Total LacCer, CE 20:3/Cer(d18:1/24:1), CE 20:3/LacCer(d18:1/20:0), CE 20:4/Cer(d18:1/18:0), CE 20:4/Cer(d18:1/24:1), CE 20:4/GlcCer(d18:1/20:0), CE 20:4/GlcCer(d18:1/24:1), CE 20:4/LacCer(d18:1/20:0), CE 20:5/LacCer(d18:1/20:0), Cer(d18:0/22:0)/Cer(d18:1/18:0), Cer(d18:0/22:0)/Cer(d18:1/20:0), Cer(d18:0/22:0)/Cer(d18:1/24:1), Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/22:0)/Total CE, Cer(d18:0/22:0)/Total DAG, Cer(d18:0/22:0)/Total GlcCer, Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/Cer(d18:1/20:0), Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/P S O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/24:0)/Total CE, Cer(d18:0/24:0)/Total Cer, DAG 16:0/18:1/Total DAG, GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), LPC 16:0/LacCer(d18:1/24:0), LPC 18:1/LacCer(d18:1/20:0), LPC 18:2/LacCer(d18:1/20:0), PC 16:0/20:3/PS O-16:0/18:2-alkenyl, PC 16:0/20:3/PS O-16:1/18:2-alkyl, PC 16:0/20:4/PS O-16:0/18:2-alkenyl, PC 16:0/20:4/PS O-16:1/18:2-alkyl, PC 16:0/20:4/Total DAG, PC 18:0/20:3/PS O-16:0/18:2-alkenyl, PC 18:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-18:2/16:0-alkenyl, PC 18:0/20:4/PS O-16:0/18:2-alkenyl, PC 18:0/20:4/PS O-16:1/18:2-alkyl, PC 18:1/18:2/PS O-16:0/18:2-alkenyl, PC 18:1/18:2/PS O-16:1/18:2-alkyl, PC 18:1/18:2/Total Cer, PC O-40:3/PS O-18:2/16:0-alkenyl, SM (d18:1/23:0) (d18:1/22:1-OH)/Total DAG, SM (d18:1/24:0) (d18:1/23:1-OH)/Total Cer, Total CE/Total DAG and Total LPC/Total LacCer.

In a preferred embodiment (for subjects that are either undergoing or not undergoing statin treatment), the one or more lipid-lipid ratio(s) whose increase is (are) compared to the control is (are) selected from (Table 9):

GlcCer(d18:1/26:1)/Total CE, Cer(d18:1/24:1)/Total PC, Cer(d18:1/24:1)/PC 16:0/20:4, Cer(d18:1/20:0)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 16:0/20:3, Total Cer/Total PC, Total LacCer/Total PC, LacCer(d18:1/20:0)/PC 18:1/18:2, PS O-16:0/18:2-alkenyl/Total PS O, Cer(d18:1/18:0)/PC 16:0/20:4, LacCer(d18:1/20:0)/Total LPC and LacCer(d18:1/20:0)/PC 16:0/20:4; In another preferred embodiment, the one or more lipid-lipid ratio(s) whose decrease is (are) compared to the control is (are) selected from (Table 9):

Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/22:0)/Cer(d18:1/24:1), DAG 16:0/18:1/Total DAG, Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/24:0)/Total CE, Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/Total Cer, Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/LacCer(d18:1/24:0), Cer(d18:0/22:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/22:0)/Cer(d18:1/20:0), Cer(d18:0/22:0)/P S O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl, GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), Total LPC/Total LacCer and GlcCer(d18:1/26:0)/LacCer(d18:1/22:0).

In a particularly preferred embodiment, the one or more lipid-lipid ratio(s) whose increase is (are) compared to the control is (are) selected from (Table 11):

Cer(d18:1/24:1)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 16:0/20:3, PS O-16:0/18:2-alkenyl/Total PS O and Cer(d18:1/18:0)/PC 16:0/20:4;

and the one or more lipid-lipid ratio(s) whose decrease is (are) compared to the control is (are) selected from (Table 11):

GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), DAG 16:0/18:1/Total DAG, Cer(d18:0/24:0)/Total Cer, Total LPC/Total LacCer, GlcCer(d18:1/26:0)/LacCer(d18:1/22:0), Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl and Cer(d18:0/24:0)/LacCer(d18:1/24:0).

In yet another embodiment the invention relates to a method of choosing an appropriate treatment of CVD and/or one or more of its complications, such as AMI or CVD death, in a subject, comprising determining in a sample from said subject one or more lipid-clinical concentration ratio(s), wherein (an) increased or decreased lipid-clinical concentration ratio(s) in said sample, when compared to a control sample, is (are) indicative of said subject being in need of treatment or a change in, or supplementation of, an already administered treatment, wherein the one or more lipid-clinical concentration ratio(s) whose increase is (are) compared to the control is (are) selected from (Tables 6a and 9a):

Cer(d18:1/16:0)/triglycerides, Cer(d18:1/18:0)/apolipoprotein A-I, Cer(d18:1/18:0)/apolipoprotein B, Cer(d18:1/18:0)/HDL cholesterol, Cer(d18:1/18:0)/total cholesterol, Cer(d18:1/18:0)/total-c/HDL-c, Cer(d18:1/18:0)/triglycerides, Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/20:0)/apolipoprotein B, Cer(d18:1/20:0)/HDL cholesterol, Cer(d18:1/20:0)/total cholesterol, Cer(d18:1/20:0)/triglycerides, Cer(d18:1/22:0)/apolipoprotein B, Cer(d18:1/22:0)/LDL cholesterol, Cer(d18:1/22:0)/triglycerides, Cer(d18:1/24:1)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein B, Cer(d18:1/24:1)/LDL cholesterol, Cer(d18:1/24:1)/total cholesterol, Cer(d18:1/24:1)/total-c/HDL-c, Cer(d18:1/24:1)/triglycerides, GlcCer(d18:1/24:0)/total cholesterol, LacCer(d18:1/16:0)/triglycerides, LacCer(d18:1/18:0)/apolipoprotein A-I, LacCer(d18:1/18:0)/apolipoprotein B, LacCer(d18:1/18:0)/HDL cholesterol, LacCer(d18:1/18:0)/LDL cholesterol, LacCer(d18:1/18:0)/total cholesterol, LacCer(d18:1/18:0)/total-c/HDL-c, LacCer(d18:1/18:0)/triglycerides, LacCer(d18:1/20:0)/apoA1/apoB, LacCer(d18:1/20:0)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein B, LacCer(d18:1/20:0)/HDL cholesterol, LacCer(d18:1/20:0)/LDL cholesterol, LacCer(d18:1/20:0)/LDL-c/HDL-c, LacCer(d18:1/20:0)/total cholesterol, LacCer(d18:1/20:0)/total-c/HDL-c, LacCer(d18:1/20:0)/triglycerides, LacCer(d18:1/22:0)/apoA/apoB, LacCer(d18:1/22:0)/apolipoprotein A-I, LacCer(d18:1/22:0)/apolipoprotein B, LacCer(d18:1/22:0)/HDL cholesterol, LacCer(d18:1/22:0)/LDL cholesterol, LacCer(d18:1/22:0)/total cholesterol, LacCer(d18:1/22:0)/total-c/HDL-c, LacCer(d18:1/22:0)/triglycerides, LacCer(d18:1/24:0)/apoA/apoB, LacCer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/24:1)/apolipoprotein B, LacCer(d18:1/24:1)/total cholesterol, LacCer(d18:1/24:1)/triglycerides, PC O-32:0 (KDdiA-PC)/apolipoprotein A-I, PC O-32:0 (KDdiA-PC)/triglycerides, PC O-34:1/triglycerides, PS O-16:0/18:1-alkenyl (PS O-16:1/18:1-alkyl)/triglycerides, PS O-16:0/18:2-alkenyl/triglycerides, PS O-16:1/18:2-alkyl/triglycerides, PS O-18:2/16:0-alkenyl/HDL cholesterol, PS O-18:2/16:0-alkenyl/triglycerides, Total Cer/apolipoprotein A-I, Total Cer/total cholesterol, Total Cer/triglycerides, Total GlcCer/apolipoprotein B, Total GlcCer/total cholesterol, Total LacCer/apolipoprotein A-I, Total LacCer/apolipoprotein B, Total LacCer/total cholesterol and Total LacCer/triglycerides;

and wherein the one or more lipid-clinical concentration ratio(s) whose decrease is (are) compared to the control is (are) selected from (Tables 6a and 9a):

CE 14:0/apolipoprotein B, CE 14:0/LDL cholesterol, CE 14:0/LDL-c/HDL-c, CE 14:0/total cholesterol, CE 14:0/total-c/HDL-c, CE 16:1/apolipoprotein B, CE 16:1/HDL cholesterol, CE 16:1/LDL cholesterol, CE 16:1/total cholesterol, CE 17:1/LDL-c/HDL-c, CE 18:3/apoA1/apoB, CE 18:3/apolipoprotein A-I, CE 18:3/apolipoprotein B, CE 18:3/HDL cholesterol, CE 18:3/LDL cholesterol, CE 18:3/LDL-c/HDL-c, CE 18:3/total cholesterol, CE 18:3/total-c/HDL-c, CE 20:3/apolipoprotein B, CE 20:3/LDL-c/HDL-c, CE 20:3/total-c/HDL-c, CE 20:5/apolipoprotein B, CE 20:5/HDL cholesterol, CE 20:5/LDL cholesterol, Cer(d18:0/22:0)/apolipoprotein B, Cer(d18:0/22:0)/LDL-c/HDL-c, Cer(d18:0/22:0)/total-c/HDL-c, Cer(d18:0/24:0)/apolipoprotein A-I, Cer(d18:0/24:0)/apolipoprotein B, Cer(d18:0/24:0)/HDL cholesterol, Cer(d18:0/24:0)/LDL cholesterol, Cer(d18:0/24:0)/LDL-c/HDL-c, Cer(d18:0/24:0)/total cholesterol, Cer(d18:0/24:0)/total-c/HDL-c, LPC 18:2/apoA1/apoB, LPC 18:2/apolipoprotein B, LPC 18:2/HDL cholesterol, LPC 18:2/LDL cholesterol, LPC 18:2/LDL-c/HDL-c, LPC 18:2/total cholesterol, PC 16:0/20:3/apolipoprotein B, PC 16:0/20:3/HDL cholesterol, PC 16:0/20:3/LDL-c/HDL-c, PC 16:0/20:3/total-c/HDL-c, PC 16:0/20:4/apolipoprotein A-I, PC 16:0/20:4/apolipoprotein B, PC 16:0/20:4/LDL cholesterol, PC 16:0/20:4/LDL-c/HDL-c, PC 16:0/20:4/total cholesterol, PC 16:0/20:4/total-c/HDL-c, PC 18:0/18:1/LDL-c/HDL-c, PC 18:0/20:3/LDL-c/HDL-c, PC 18:0/20:3/total-c/HDL-c, PC 18:0/20:4/apoA1/apoB, Total LPC/LDL-c/HDL-c, Total LPC/total-c/HDL-c, Total PC/apolipoprotein B, Total PC/LDL-c/HDL-c, Total PC/total cholesterol and Total PC/total-c/HDL-c.

In yet another embodiment the invention relates to a method of choosing an appropriate treatment of CVD and/or one or more of its complications, such as AMI or CVD death, in a subject, wherein the subject is not undergoing statin treatment and wherein said method comprises determining in a sample from said subject one or more lipid-clinical concentration ratio(s), wherein (an) increased or decreased lipid-clinical concentration ratio(s) in said sample, when compared to a control sample, is (are) indicative of said subject being in need of treatment or a change in, or supplementation of, an already administered treatment, wherein the one or more lipid-clinical concentration ratio(s) whose increase is (are) compared to the control is (are) selected from (Tables 6b and 9b):

Cer(d18:1/16:0)/apolipoprotein A-I, Cer(d18:1/16:0)/LDL cholesterol, Cer(d18:1/16:0)/triglycerides, Cer(d18:1/18:0)/apolipoprotein A-I, Cer(d18:1/18:0)/apolipoprotein B, Cer(d18:1/18:0)/HDL cholesterol, Cer(d18:1/18:0)/total cholesterol, Cer(d18:1/18:0)/total-c/HDL-c, Cer(d18:1/18:0)/triglycerides, Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/20:0)/apolipoprotein B, Cer(d18:1/20:0)/HDL cholesterol, Cer(d18:1/20:0)/LDL cholesterol, Cer(d18:1/20:0)/total cholesterol, Cer(d18:1/20:0)/total-c/HDL-c, Cer(d18:1/20:0)/triglycerides, Cer(d18:1/22:0)/apolipoprotein A-I, Cer(d18:1/22:0)/apolipoprotein B, Cer(d18:1/22:0)/LDL cholesterol, Cer(d18:1/22:0)/total cholesterol, Cer(d18:1/22:0)/triglycerides, Cer(d18:1/24:0)/apolipoprotein A-I, Cer(d18:1/24:0)/apolipoprotein B, Cer(d18:1/24:0)/LDL cholesterol, Cer(d18:1/24:0)/total cholesterol, Cer(d18:1/24:1)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein B, Cer(d18:1/24:1)/HDL cholesterol, Cer(d18:1/24:1)/LDL cholesterol, Cer(d18:1/24:1)/LDL-c/HDL-c, Cer(d18:1/24:1)/total cholesterol, Cer(d18:1/24:1)/total-c/HDL-c, Cer(d18:1/24:1)/triglycerides, GlcCer(d18:1/20:0)/apolipoprotein B, GlcCer(d18:1/20:0)/total cholesterol, GlcCer(d18:1/24:1)/apolipoprotein B, GlcCer(d18:1/26:1)/apolipoprotein A-I, LacCer(d18:1/16:0)/triglycerides, LacCer(d18:1/18:0)/apolipoprotein A-I, LacCer(d18:1/18:0)/apolipoprotein B, LacCer(d18:1/18:0)/total cholesterol, LacCer(d18:1/20:0)/apoA1/apoB, LacCer(d18:1/20:0)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein B, LacCer(d18:1/20:0)/HDL cholesterol, LacCer(d18:1/20:0)/LDL cholesterol, LacCer(d18:1/20:0)/LDL-c/HDL-c, LacCer(d18:1/20:0)/total cholesterol, LacCer(d18:1/20:0)/total-c/HDL-c, LacCer(d18:1/20:0)/triglycerides, LacCer(d18:1/22:0)/apoA1/apoB, LacCer(d18:1/22:0)/apolipoprotein A-I, LacCer(d18:1/22:0)/apolipoprotein B, LacCer(d18:1/22:0)/HDL cholesterol, LacCer(d18:1/22:0)/LDL cholesterol, LacCer(d18:1/22:0)/total cholesterol, LacCer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/24:1)/apolipoprotein B, LacCer(d18:1/24:1)/total cholesterol, LacCer(d18:1/24:1)/triglycerides, PC O-34:1/apolipoprotein B, PS O-16:0/18:2-alkenyl/triglycerides, PS O-16:1/18:2-alkyl/triglycerides, Total Cer/apolipoprotein A-I, Total Cer/apolipoprotein B, Total Cer/LDL cholesterol, Total Cer/total cholesterol, Total DAG/apolipoprotein A-I, Total DAG/triglycerides, Total GlcCer/apolipoprotein B, Total LacCer/apolipoprotein A-I, Total LacCer/apolipoprotein B and Total LacCer/total cholesterol.

and wherein the one or more lipid-clinical concentration ratio(s) whose decrease is (are) compared to the control is (are) selected from (Tables 6b and 9b):

CE 14:0/apoA1/apoB, CE 14:0/apolipoprotein B, CE 14:0/LDL-c/HDL-c, CE 14:0/total-c/HDL-c, CE 16:1/apoA1/apoB, CE 18:3/apoA1/apoB, CE 18:3/apolipoprotein A-I, CE 18:3/apolipoprotein B, CE 18:3/HDL cholesterol, CE 18:3/LDL cholesterol, CE 18:3/LDL-c/HDL-c, CE 18:3/total cholesterol, CE 18:3/total-c/HDL-c, CE 20:5/triglycerides, Cer(d18:0/24:0)/apolipoprotein B, Cer(d18:0/24:0)/total cholesterol, Cer(d18:0/24:0)/total-c/HDL-c, PC 18:0/20:4/apoA1/apoB, PC O-38:6/apolipoprotein A-I, Total LPC/apoA1/apoB and Total PC/apolipoprotein A-I.

In a preferred embodiment (for subjects that are either undergoing or not undergoing statin treatment), the one or more lipid-clinical concentration ratio(s) whose increase is (are) compared to the control is (are) selected from (Table 10):

Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein A-I, Total Cer/apolipoprotein A-I, Total LacCer/apolipoprotein A-I, Cer(d18:1/18:0)/apolipoprotein A-I, LacCer(d18:1/22:0)/apolipoprotein A-I, LacCer(d18:1/20:0)/HDL cholesterol and Cer(d18:1/24:1)/apolipoprotein B.

In another preferred embodiment, the one or more lipid-clinical concentration ratio(s) whose decrease is (are) compared to the control is (are) selected from (Table 10):

Cer(d18:0/24:0)/apolipoprotein B, Cer(d18:0/24:0)/total cholesterol, Cer(d18:0/24:0)/apolipoprotein B, PC 16:0/20:4/apolipoprotein B and Cer(d18:0/24:0)/apolipoprotein A-I.

In a particularly preferred embodiment, the one or more lipid-clinical concentration ratio(s) whose increase is (are) compared to the control is (are) selected from (Table 11):

Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein A-I, Total LacCer/apolipoprotein A-I, LacCer(d18:1/20:0)/HDL cholesterol and Cer(d18:1/18:0)/apolipoprotein A-I.

For the purposes of the invention, and particularly for lipid-clinical concentration ratios, an Apolipoprotein A-I measurement may alternatively be an Apolipoprotein A-II measurement.

In one embodiment of the invention, the treatment the effectiveness of which is to be evaluated or which is to be chosen as appropriate in accordance with the methods described and claimed herein, is a lipid modifying treatment.

For the purposes of the invention, at least one lipid concentration, lipid-lipid ratio or lipid-clinical concentration ratio from Tables 4-13, or combinations thereof, may be determined to assess whether the patient is at risk to develop one or more of CVD complications such as AMI or CVD death; to evaluate the effectiveness of the treatment of CVD and/or one or more of its complications, such as AMI or CVD death in a subject; or to choose an appropriate treatment of CVD and/or one or more of its complications, such as AMI or CVD death in a subject. However, it is also possible, and may be advantageous, to determine at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, or at least 8 lipid concentrations, lipid-lipid ratios or lipid-clinical concentration ratios from Tables 4-13, or combinations thereof, in this regard. Where more than one lipidomic markers are determined and used for the assessment, it may be advantageous that a specific lipid concentration, lipid-lipid ratio, lipid-clinical concentration ratio or combination thereof, is given greater weight than others in the above-mentioned assessment, evaluation or choice.

Preferred embodiments of the invention are methods wherein the one or more lipid(s) or lipid ratio(s), or combination thereof, comprise(s): Cer(d18:1/20:0), LacCer(d18:1/20:0), Cer(d18:1/24:1), LacCer(d18:1/24:1), LacCer(d18:1/22:0), Cer(d18:1/18:0), PC 16:0/20:4, Cer(d18:0/24:0), Cer(d18:1/24:1)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 16:0/20:3, PS O-16:0/18:2-alkenyl/Total PS O, Cer(d18:1/18:0)/PC 16:0/20:4, GlcCer(d18:1/26:0)/LacCer(d18:1/20:0), DAG 16:0/18:1/Total DAG, Cer(d18:0/24:0)/Total Cer, Total LPC/Total LacCer, GlcCer(d18:1/26:0)/LacCer(d18:1/22:0), Cer(d18:0/24:0)/Cer(d18:1/24:1), Cer(d18:0/24:0)/Cer(d18:1/18:0), Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/24:0)/Cer(d18:1/22:0), Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl, Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl, Cer(d18:0/24:0)/LacCer(d18:1/24:0), Cer(d18:1/20:0)/apolipoprotein A-I, Cer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein A-I, Total LacCer/apolipoprotein A-I, LacCer(d18:1/20:0)/HDL cholesterol and Cer(d18:1/18:0)/apolipoprotein A-I.

In the context of the present invention, CVD is typically characterized by coronary artery disease, peripheral artery disease, a stroke and/or CVD death. The CVD in the subject whose sample is analyzed in accordance with the invention may be atherosclerosis-induced. However, the invention also embodies methods involving subjects who are at risk of developing CVD, but who may or may not have atherosclerosis.

In a further embodiment, the methods of the invention may further comprise determining the serum level of total cholesterol, low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), Apolipoprotein B (ApoB) and/or Apolipoprotein C-III in the subject's sample. In one embodiment of the invention, the subject does not have elevated serum levels of one or more of total cholesterol, low-density lipoprotein cholesterol (LDL-C), Apolipoprotein C-III or Apolipoprotein B (ApoB), or a decreased serum level of HDL-cholesterol (HDL-C).

As mentioned above, for the purposes of the present invention, a control sample may be obtained from (a) CAD patient(s) or a group of CAD patients that has/have remained free of any major CVD complications e.g., by mixing a variety of samples from said population. If a group of CAD patients is used then several lipid profiles from a population are combined and the lipidomic marker is created from this combination. The levels or amounts of the individual lipids or the lipid-lipid ratios or lipid-clinical concentration ratios in the sample from a subject are compared to the levels or amounts of the lipids or lipid ratios in the control for determining the risk of one or more of CVD complications, such as AMI or CVD death, in said subject.

The invention encompasses the analysis of lipid concentrations, lipid-lipid ratios and/or lipid-clinical concentration ratios in samples from a subject that has been or is being treated with one or more statins and/or any other HMG-CoA reductase inhibitor.

Alternatively, the invention encompasses the analysis of lipid concentrations, lipid-lipid ratios and/or lipid-clinical concentration ratios in samples from a subject that has not yet undergone statin therapy or therapy with any other HMG-CoA reductase inhibitor.

In accordance with the aspects and embodiments of the invention described and claimed herein, the statin may be one selected from the group consisting of atorvastatin, cerivastatin, fluvastatin, fluvastatin XL, lovastatin, pitavastatin, pravastatin, rosuvastatin, and simvastatin.

Collecting information on a lipidomic marker or a lipidomic profile from a subject's biological sample can be performed via various chemical and high resolution analytical techniques. Suitable analytical techniques include but are not limited to mass spectrometry and nuclear resonance spectroscopy. Any high resolution technique capable of resolving individual lipids or lipid classes and providing structural information of the same can be used to collect the lipid profile from the biological sample. For methods of the present invention the level of the lipid is determined by using mass spectrometry, nuclear magnetic resonance spectroscopy, fluorescence spectroscopy or dual polarisation interferometry, a high performance separation method such as HPLC or UPLC and/or an immunoassay such as an ELISA. According to an alternative or further embodiment an analyte in a sample can be detected and/or quantified by combining the analyte with a binding moiety capable of specifically binding the analyte. The binding moiety can include, for example, a member of a ligand-receptor pair, i.e., a pair of molecules capable of having a specific binding interaction. The binding moiety can also include, for example, a member of a specific binding pair, such as antibody-antigen, enzyme-substrate, nucleic acid-based ligands, other protein ligands, or other specific binding pairs known in the art. In a preferred embodiment, the lipidomic profile is collected with mass spectrometry (MS), wherein the MS instrument may be coupled to direct infusion methods and high performance separation methods such as HPLC or HPLC. The amount of the individual lipids or lipid classes in the collected lipidomic profile is used when comparing the collected lipid profile to a control.

The methods of the present invention may be used for determining a risk of said patient to develop CVD complications, particularly severe CVD complications such as death and myocardial infarction (MI), including acute myocardial infarction (AMI).

In one embodiment of the invention, a method for treating or preventing CVD complications, such as AMI or CVD death, in a subject in need thereof is provided. The method comprises administering a therapeutically effective dose of a drug capable of modulating one or more of the lipid concentration(s), lipid-lipid ratio(s) or lipid-clinical concentration ratio(s) described in Tables 4-13, wherein the dose is such that said one or more lipid concentration(s), lipid-lipid ratio(s) or lipid-clinical concentration ratio(s) in a sample of said subject does not significantly change when compared to (a) corresponding lipid concentration(s), (a) corresponding lipid-lipid ratio(s) or (a) corresponding lipid-clinical concentration ratio(s) in a control, e.g., a control sample. In a preferred embodiment, the drug is a statin or another HMG CoA reductase inhibitor. Particularly preferred statins in this regard are atorvastatin, cerivastatin, fluvastatin, fluvastatin XL, lovastatin, pitavastatin, pravastatin, rosuvastatin or simvastatin. In another preferred embodiment, the drug is niacin (nicotinic acid); a cholesterol absorption inhibitor, such as ezetimibe or SCH-48461; a cholesteryl ester transfer protein (CETP) inhibitor, such as torcetrapib, anacetrapib or JTT-705; a bile acids sequestrant, such as colesevelam, cholestyramine and colestipol; or a fibrate, such as fenofibrate, gemfibrozil, clofibrate, and bezafibrate. Alternatively, it may also be a phytosterol.

Also embodied by the present invention is a lipid as described herein, e.g. a lipid from any of Tables 4, 7, 10 or 13, for use in preventing or treating a subject at risk to develop CVD complications such as AMI or CVD death, wherein the said lipid is to be taken as a dietary supplement or a medicament. A corresponding method of treatment is likewise encompassed. Likewise, the invention also encompasses a modulator for use for modulating a lipid concentration, lipid-lipid ratio or lipid-clinical concentration ratio as described herein, e.g., in Tables 4-13, in a subject at risk to develop CVD and/or one or more of its complications such as AMI or CVD death. A corresponding method of treatment is likewise encompassed. In a further embodiment, the said modulator is a small molecule, an antisense RNA, a small interfering RNA (siRNA) or a natural or modified lipid.

In one embodiment of the present invention, an antibody against any one of the lipids in Tables 4-13 is used for predicting one or more CVD complications such as AMI or CVD death. In another embodiment of the invention, the antibody may be used for preventing or treating one or more of the above complications in a subject.

Any of the methods, drugs, lipids or antibodies of the present invention may be used for a subject which is at risk to develop or has suffered from one or more CVD complications such as acute myocardial infarction and/or a cardiovascular death. For the purposes of the invention, CVD complication(s) includes severe CVD complication(s), particularly death.

Also encompassed by the present invention is a kit for predicting CVD complications or for performing any of the methods or uses of the present invention, wherein the kit comprises a lipid standard chosen from the lipids in Tables 4, 7, 10 or 13, one or more control lipidomic markers, an antibody against one of the said lipids, and reagents for performing the method.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1.

This FIGURE demonstrates the importance of molecular lipid measurements. Two examples are given in this FIGURE to illustrate that two closely related molecular lipids may have a very different or even opposite effect on CVD complications. First quadrangles are used to indicate two Lactosylceramides (LacCer). LacCer (18:1/20:0) is a significant predictor for CVD Death, while closely related LacCer (18:1/16:0) has only limited or no value as a risk predictor. Second example proves that that two lipid species from the same lipid class may have even an opposite effect on CVD events. PC (18:0/20:4) proved to be a protective lipid against CVD death while PC (16:0/16:0) seems to increase CVD complications.

DETAILED DESCRIPTION OF THE INVENTION

Definitions:

Coronary vascular disease/cardiovascular disease (CVD) has its general meaning in the art and is used to classify numerous conditions that affect the heart, heart valves, blood, and vasculature of the body, including CAD. In the present invention the terms CVD and CAD may be used interchangeably. Cardiovascular diseases include endothelial dysfunction, coronary artery disease, angina pectoris, myocardial infarction, atherosclerosis, congestive heart failure, hypertension, cerebrovascular disease, stroke, transient ischemic attacks, deep vein thrombosis, peripheral artery disease, cardiomyopathy, arrhythmias, aortic stenosis, and aneurysm. Such diseases frequently involve atherosclerosis. In a preferred embodiment of the invention, the cardiovascular disease is a cardiovascular disease associated with atherosclerosis.

CAD is coronary artery disease, AMI is acute myocardial infarction, ACS is acute coronary syndrome, CAC is coronary artery calcification, RCT is reverse cholesterol transport, LDL is low density lipoprotein, HDL is high density lipoprotein, LDL-C is low density lipoprotein cholesterol, HDL-C is high density lipoprotein cholesterol, ApoA is Apolipoprotein A, ApoB is Apolipoprotein B, ApoC is apolipoprotein C, MS is mass spectrometry, HPLC is high performance liquid chromatography, and UPLC is ultra performance liquid chromatography.

As used herein, “a subject” includes all mammals, including without limitation humans, but also non-human primates, dogs, cats, horses, sheep, goats, cows, rabbits, pigs and rodents.

A “sample” is defined as any biological sample obtained from a subject or a group or population of subjects. For the purposes of the present invention, the biological sample may be whole blood, blood serum, or blood plasma. It may also be a tissue sample. However, a preferred embodiment is wherein the biological sample is plasma or serum. Taking a blood sample of a patient is a part of normal clinical practice. The blood sample can be taken in connection with e.g. measuring the cholesterol levels in the patients. The collected blood sample can be prepared and serum or plasma can be separated with techniques well known to a person skilled in the art. Vena blood samples can be collected from patients using a needle and a BD Vacutainer® Plastic Tubes or Vacutainer® Plus Plastic Tubes (BD Vacutainer® SST™ Tubes contain spray-coated silia and a polymer gel for serum separation). Serum can be separated by centrifugation at 1300 RCF for 10 min at room temperature and stored in small plastic tubes at −80° C.

For the purposes of the present invention, lipids from the Lipidomic analysis were named according to the following nomenclature: CE is cholesteryl ester, Cer is ceramide, DAG is diacylglycerol, PC O is ether-linked PC, GD is disialogangliosides, GlcCer is galactosyl- and glucosylceramides, GM is monosialogangliosides, LacCer is lactosylceramides, LPC is lysophosphatidylcholine, PC is Phosphatidylcholine, PE is Phosphatidylethanolamine, PI is Phosphatidylinositol, SM is Sphingomyelin, S1P is sphingosine-1-phosphate.

The nomenclature X:Y indicates, X number of total carbon atoms in the fatty acid(s) portions of the molecule, and Y the total number of double bonds in the fatty acid portion(s) of the molecule.

The nomenclature A/B indicates, for a molecule of DAG and PC, A and B types of fatty acid moieties attached to the glycerol backbone of the molecule.

The nomenclature (dC/A) indicates, for a molecule of Cer, GlcCer, LacCer and SM, C the type of long-chain base with an amide-linked, A, fatty acid moiety.

The wording “compared to a control sample” as used herein will be understood to include embodiments where control samples are actually analyzed in respect of a lipidomic marker of interest, i.e., in respect of the concentration of one or more of the lipid(s), the lipid-lipid ratios, or the lipid-clinical concentration ratios or combinations thereof as specifically described herein in connection with the various aspects and embodiments of the present invention. It will be appreciated, however, that the above wording also includes embodiments where the corresponding information on said lipidomic marker in said control sample is merely taken from the literature, or has been previously determined, calculated or extrapolated, or is yet to be determined, calculated or extrapolated.

As used herein, the term “antibody” includes monoclonal and polyclonal antibodies, whole antibodies, antibody fragments, and antibody sub-fragments that exhibit specific binding to a said lipid. Thus, suitable “antibodies” can be whole immunoglobulins of any class, e.g., IgG, IgM, IgA, IgD, IgE, chimeric antibodies or hybrid antibodies with dual or multiple antigen or epitope specificities, or fragments, e.g., F(ab′)₂, Fab′, Fab and the like, including hybrid fragments, and additionally includes any immunoglobulin or any natural, synthetic or genetically engineered protein that acts like an antibody by binding to a specific antigen to form a complex. The term “antibody” encompasses antigen-binding fragments of antibodies (e.g., single chain antibodies, Fab fragments, F(ab′)₂, a Fd fragment, a Fv fragment and dAb fragments) as well as complete antibodies. For example, Fab molecules can be expressed and assembled in a genetically transformed host like E. coli. A lambda vector system is available thus to express a population of Fab's with a potential diversity equal to or exceeding that of subject generating the predecessor antibody. See Huse W D, et al., Science 1989, 246:1275-81. Such Fab's are included in the definition of “antibody.” The ability of a given molecule, including an antibody fragment or sub-fragment, to act like an antibody and specifically bind to a specific antigen can be determined by binding assays known in the art, for example, using the antigen of interest as the binding partner.

Antibodies against lipids in accordance with the present invention may be prepared by methods well known to those skilled in the art. For example, mice may be immunized with a lipid with adjuvant. Splenocytes are harvested as a pool from the mice that were administered 3 immunizations at 2-week intervals with test bleeds performed on alternate weeks for serum antibody titers. Splenocytes are prepared as 3 aliquots that are either used immediately in fusion experiments or stored in liquid nitrogen for use in future fusions.

Fusion experiments are then performed according to the procedure of Stewart & Fuller, J. Immunol. Methods 1989, 123:45-53. Supernatants from wells with growing hybrids are screened by enzyme-linked immunosorbent assay (ELISA) for monoclonal antibody (MAb) secretors on 96-well ELISA plates coated with the said lipid. ELISA positive cultures are cloned by limiting dilutions, typically resulting in hybridomas established from single colonies after 2 serial cloning experiments.

EXAMPLES Example 1

Materials and Methods

This study is a sub-cohort of the LURIC study that is a large scale prospective study on cardiovascular epidemiology. LURIC database contains clinical information over 3000 patients including baseline coronary angiography, clinically used biomarker data and also e.g. CVD mortality data for the follow-up period (3 years). In this biomarker study the inventors compared CAD cases (n=62) that died during the follow-up due to CVD with patients (n=173) having a stable CAD. Subjects with a significant atherosclerosis level in the angiogram but no CVD related death during the follow-up were used as controls, while the case group had similarly a significant atherosclerosis based on the angiography at baseline and in addition they died during the follow-up due to acute cardiovascular events. A statistical analysis was performed separately also for cases (n=48) and controls (n=124) that were not treated with statins. The clinical characteristics are described in Table 1.

TABLE 1 Background characteristics for LURIC patients analyzed with lipidomics Variable Controls (n = 173) Cases (n = 62) Age (average) 60 67 LDL-C (mg/dL) 122 117.5 HDL-C (mg/dL) 37.2 35.3 DM2 patients 62 (36%) 36 (58%) Hypertensive patients 101 (58%)  39 (63%) Lipid lowering users 49 (28%) 14 (23%) Smokers (active or quit less than 3 46 (27%)  8 (13%) years before sampling)

Definition of Cases: All cases had a significant vessel disease (>=20% stenosis) in coronary angiogram and they all died due to CVD during the follow-up.

Definition of Controls: All controls had a significant vessel disease (>=20% stenosis) in coronary angiogram, but they did not die due to CVD during the follow-up.

Example 2

Analytical Methods

Mass Spectrometry Driven Lipidomics

Direct infusion coupled to tandem mass spectrometry, i.e. shotgun lipidomics, and two liquid chromatography tandem mass spectrometry (LC-MS/MS) approaches, i.e. ceramide and cerebroside lipidomics, were used to identify lipid biomarkers for coronary artery disease (CVD) risk by analyzing molecular lipid species in human serum, plasma, and carotid artery plaques. The applied methods were optimized especially for quantification of molecular cholesteryl esters (CE), phosphatidylcholines (PC), lysophosphatidylcholines (LPC) and other lysophospholipids (LPL), ether-linked phosphatidylcholines (PC O) and other ether-linked phospholipids (PL O), phosphatidylserines (PS), phosphatidylethanolamines (PE), phosphatidylglycerols (PG), phosphatidylinositols (PI), phosphatidic acids (PA), diacylglycerols (DAG), ceramides (Cer), glucosylceramides (GlcCer) and lactosylceramides (LacCer).

The following materials were used according to the methods. HPLC or LC-MS grade of chloroform, methanol, water, acetonitrile, formic acid, methanol, isopropanol, ammonium acetate, acetic acid, potassium chloride and butylated hydroxytoluene (BHT) were purchased from Sigma-Aldrich (St. Louis, Mo., USA).

HPLC column (Acquity BEH C18, 2.1×50 mm id. 1.7 m) was purchased from Waters (Milford, Mass., USA). HPLC pre-column (Widepore C18 4×2.0 mm) was purchased from Phenomenex (Torrance, Calif., USA). All labware used for the extraction were resistant to chloroform. Aerosol resistant filter tips (Molecular BioProducts) and Eppendorf 2 ml safe-lock tubes, 96-well twin.tec PCR plates, and Pierce-it-lite thermo-sealing foils were purchased from VWR International (West Chester, Pa., USA). CO-RE Filter Tips and 96-well 2 ml Whatman Uniplates were purchased from Hamilton Robotics (Bonaduz, Switzerland). Synthetic lipid standards were purchased from Avanti Polar Lipids (Alabaster, Ala., USA) and from Matreya (Pleasant Gap, Pa., USA).

Lipids were extracted in chloroform:methanol according to the following protocols. Samples were spiked with known amounts of non-endogenous synthetic internal standards for data normalization and endogenous lipid quantification. In shotgun lipidomics analysis; LPC 17:0, PC 17:0/17:0, PA 17:0/17:0, PE 17:0/17:0, PG 17:0/17:0, PS 17:0/17:0, DAG 17:0/17:0, D6-CE 18:0, in ceramide and cerebroside lipidomics; Cer d18:1/17:0, D3-LacCer d18:1/16:0, and D3-GlcCer d18:1/16:0, were used as internal standards. Post-extract spiked non-endogenous synthetic external standards were used for quality controlling. Stock solutions of standards were prepared by dissolving appropriately weighed amounts of each standard in chloroform:methanol (2:1, V:V) to achieve a final concentration of 500 μM. An internal standard mixture containing each of the standard stock was created and used in lipid extraction.

Samples and quality control samples for each extraction batch were thawed on ice. The carotid artery plaque samples were weighed on ice by using a cryo-box and homogenized in ice-cold 70% methanol in water. The Mixer Mill 301 Teflon adapters were kept at −20° C. Homogenization was performed at 15-25 Hz for 2-15 minutes with Mixer Mill 301 (Retch GmbH, Germany).

Lipid extraction of human samples was carried out in automated fashion using a Hamilton MICROLAB STAR system (Hamilton Robotics, Switzerland). Well-mixed samples were aliquoted into a 96-well 2 ml Whatman Uniplate containing ice-cold methanol and 0.1% BHT. 5 μl of serum and plasma and 30 μl of carotid artery plaques were used for shotgun- and ceramide and cerebroside lipidomics and 100 μl of serum and plasma. The samples were mixed thoroughly after each step in the extraction protocol. The extraction proceeded at room temperature by adding an appropriate volume of internal standard mixture and chloroform. In shotgun and ceramide and cerebroside lipidomics, the organic phase separation was facilitated by adding 20 mM acetic acid and centrifuging the plate for 5 min at 500×g. The organic phase was transferred into a new 96-well 2 ml Whatman Uniplate. The remaining water-containing phase was washed by adding appropriate volume of chloroform followed by centrifugation. The two organic phases were pooled and evaporated under N₂ until dryness. The lipid extracts were then re-dissolved in chloroform:methanol (1:2, v:v) including the addition of the synthetic external standard.

In shotgun lipidomics, lipid extracts were analyzed on a hybrid triple quadrupole/linear ion trap mass spectrometer (QTRAP 5500, AB Sciex) equipped with a robotic nanoflow ion source (NanoMate HD, Advion Biosciences). The instruments were operated in positive and negative ion modes. In positive ion the spray voltage was set to 1.0 to 1.4 kV and in negative ion mode to −1.0 to −1.4 kV. A gas pressure of 0.3-0.8 psi was used and the interface heater was set at 60° C. The collision energy (CE) and declustering potential (DP) was optimized for each lipid class using synthetic standards. The mass spectrometer was operated in unit resolution mode using a scan speed of 200 Da/s. Molecular lipids were analyzed in both positive and negative ion modes using multiple precursor ion scanning (MPIS) and neutral loss scanning (NLS) as described by Stahlman and colleagues (Stahlman M, et al. High-throughput shotgun lipidomics by quadrupole time-of-flight mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci 2009).

In ceramide and cerebroside lipidomics, the high performance liquid chromatography (HPLC) analyses were conducted in the following way. Chromatographic apparatus consisted of a CTC HTC PAL autosampler (CTC Analytics AG, Switzerland), a Rheos Allegro UHPLC pump (Flux Instruments AG, Switzerland), an external column heater set to 60° C. for ceramide and cerebroside lipidomics. The extracted samples, 10 μl of each, were injected into the pre-column followed by the analytical column and delivered to the mass spectrometer at a flow rate of 500 μl/min. In ceramide and cerebroside lipidomics, A gradient was used for lipid analyte separation with solvent A comprising 10 mM ammonium acetate in HPLC grade water containing 0.1% formic acid and solvent B of 10 mM ammonium acetate in acetonitrile:isopropanol (4:3, V:V) containing 0.1% formic acid. The gradient was constructed in the following way: 0 min-65% B; 2 min-65% B; 2.5 min-75% B; 17.5 min-100% B; 22.5 min-100% B; 22.6 min-65% B; 25 min-65% B.

The lipid extracts were analyzed by HPLC-MS/MS. The MS analysis was performed on a hybrid triple quadrupole/linear ion trap mass spectrometer equipped with the Turbo V™ Ion Source (4000 QTRAP, AB Sciex). The instrument was operating in positive and negative ion modes. The ion source voltage was set to 5500V for ceramide and cerebroside lipidomics. The collision energy (CE) and declustering potential (DP) was optimized for each lipid class using synthetic standards. A 20 sec dwell time was applied for each scan.

The data processing was done in the following way. Initially the retention time (in LC mode) and identification of each peak was done using endogenous standards and by Information Dependent Acquisition (IDA) experiments where applicable. The raw data were processed according to peak detected and retention time (in LC mode) in automated fashion. A stringent cutoff was applied for separating background noise from actual lipid peaks. Each sample was controlled and only accepted when fulfilling the stringent acceptance criteria. Peak area counts (cps) of detected peaks were converted into a list of corresponding lipid names. Lipids were normalized to their respective internal standard and sample volume or tissue weight to retrieve their concentrations.

Several quality controls were used in the lipidomic analyses. A calibration line using synthetic or isolated standards was obtained prior to sample analysis. Synthetic standards were chosen based on application and had similar properties to the endogenous lipids or analyte(s) of interest. The calibration line consisted of a minimum of five standards points covering the expected quantification range. A sample extracted without standard and standards extracted with no matrix, were included with the calibration line.

The calibration line was used to determine the dynamic quantification range for each lipid class monitored, e.g., the linear quantification limits. As the internal standards used behave in the same way as endogenous lipids they were used for quantifying endogenous lipid species. The calibration lines were based on the same internal standards that were used for quantification of the endogenous lipids.

In each sample extracted for lipids, the ratio of synthetic internal standards (IS) to corresponding post-extract spiked external standard (ES) was determined. The peak area (cps) ratio of internal to external standard (IS/ES) was used for calculating the Coefficient of Variation (CV) across all samples. The IS/ES ratio enabled the calculation of lipid extraction recovery.

Instrument control (IC) was included at the start, middle and end of each run. IC sample analyzed was an extracted reference plasma sample and a set of standards to monitor the instrument's performance, i.e., the intra- and inter-assay variation.

For each platform, a stringent cutoff was applied for separating background noise from actual lipid peaks. Each sample was controlled and only accepted when fulfilling the stringent acceptance criteria. Masses and counts of detected peaks were converted into a list of corresponding lipid names. Lipids were normalized to their respective internal standard and sample volume to retrieve their concentrations.

Statistical Analyses

Percentage changes in lipid concentrations between control and case groups were calculated as follows: 100*(AVG[C] in case group−AVG[C] in control group)/AVG[C] in control group

Statistical significance was assigned based on standard t-test p-values.

In addition, ROC curves were used for finding lipid molecules and concentration cutoffs that separate the best cases from controls. Selectivity is calculated as a number of correctly identified cases divided by the total number of cases. Specificity is calculated as a number of correctly identified controls divided by the total number of controls. Selectivity and specificity was calculated for each lipid concentration, lipid to lipid ratio and ratio of lipid to clinical concentrations.

Example 3

Ethics

The LURIC study was approved by the ethics review committee at the “Landesärztekammer Rheinland-Pfalz” (Mainz, Germany). Written informed consent was obtained from each of the participants.

Results

In this LURIC study sub-cohort, the traditional biomarkers including LDL-cholesterol and HDL-cholesterol concentrations were practically identical in both groups and therefore were not predictive of CVD-related mortality in this study.

Multiple lipidomic markers appeared as significant predictors of CVD death (Tables 4-13). A total of 151 molecular lipids were quantified. The significant predictors were selected based on the top fifty candidates from each category, when available. The biomarker candidates based on molecular lipid concentrations are presented in Tables 4, 7, 10 and 13. The candidates were selected according to the following criteria: t-test p-value≤0.05 or sensitivity≥60% and specificity≥60%. From traditional clinical chemistry only apolipoprotein A1 and total cholesterol reached statistical significance with p-value lower than 0.05, but % change was less than 10% between controls and cases, other clinical values did not show any statistical significance. The predictive value of new lipidomic biomarkers was increased when their levels were expressed as distinct lipid-lipid ratios or lipid-clinical ratios (e.g. LDL-C or HDL-C). The top biomarker candidates are presented in Table 11. Top candidates from each category, when available, were selected based on the following selection criteria: t-test p-value≤0.05 and sensitivity≥60% and specificity≥60%.

Importance of Detailed Molecular Lipid Analyses

Recent evolvement of mass spectrometry driven lipid analysis approaches has made it possible to resolve complex lipidomes to their molecular lipid species level at high-throughput and quality required for analyses of clinical cohorts. As a result of the high sensitivity and selectivity of the methods, a lipidome-wide analysis of minute sample amounts has become feasible. Present technologies are capable of identifying lipids with different sum compositions, i.e. phosphatidylcholine (PC) 34:1, but more important is the identification of molecular lipid species, e.g. PC 16:0/18:1. In the latter analysis, information of the type of fatty acids and their positions attached to the glycerol backbone making up the particular PC molecule is retrieved.

The seminal work of Shinzawa-Itoh and colleagues showed by highly sophisticated experiments that the oxygen transfer mechanism in cytochrome c oxidase requires a specific phosphatidylglycerol molecular lipid with palmitate and vaccenate at the sn-1 and sn-2 positions respectively on the glycerol backbone (Shinzawa-Itoh K, Aoyama H, Muramoto K et al: Structures and physiological roles of 13 integral lipids of bovine heart cytochrome c oxidase. EMBO J 2007, 26:1713-1725). In line with other studies, this undoubtedly indicates that the lipid structure is an essential determinant of the biological effect. Therefore, molecular lipidomics is an essential for biomarker discovery. FIG. 1 illustrates the importance of molecular lipid data by comparing the biomarker value of two PC and LacCer molecules in predicting CVD mortality in the LURIC cohort. The data reveals that while LacCer(d18:1/20:0) is a significant CVD predictor, LacCer (d18:1/18:16:0) has low biomarker potential. In addition, two PC molecules PC (18:0/20:4) and PC (18:0/16:0) have even opposite effects on CVD complications. Thus, it is always necessary to identify and quantify all lipid species for lipid classes of interest including but not limited to cholesterol esters, different phospholipid classes, ceramides, cerebrosides (lactosylceramides, glycosylceramides), and gangliosides.

TABLE 2 Significant lipids in LURIC study sorted by p-value. Lipid names, p-values and % change for negative correlation are presented. Table 2a shows significant lipids from all study subjects. The significant lipids from subjects not undergoing statin treatment are listed in table 2b. Lipid name p-value Percentage change 2a) Significant lipids in LURIC study sorted by p-value from all study subjects. Positive correlation LacCer(d18:1/20:0) 0.00008 29.52421 LacCer(d18:1/22:0) 0.00046 22.75541 LacCer(d18:1/18:0) 0.00094 26.78692 Cer(d18:1/18:0) 0.00177 23.30373 Cer(d18:1/20:0) 0.00302 17.32385 LacCer(d18:1/24:1) 0.00361 24.72456 PS O-18:2/16:0-alkenyl 0.00571 49.89286 PS O-16:0/18:2-alkenyl 0.00670 49.99084 PS O-16:1/18:2-alkyl 0.00670 49.99084 Cer(d18:1/24:1) 0.01142 14.88898 Total LacCer 0.01669 13.83938 PS O-18:0/18:2-alkenyl 0.02899 40.93773 (PS O-18:1/18:2-alkyl) LacCer(d18:1/24:0) 0.04425 15.80958 PC O-32:0 (KDdiA-PC) 0.04546 17.40815 GlcCer(d18:1/18:0) 0.04913 12.83156 Negative correlation Total PC 0.00011 −16.07367 PC 16:0/20:4 0.00077 −18.06547 Total LPC 0.00126 −17.02070 CE 14:0 0.00181 −22.73309 PC 16:0/20:3 0.00191 −18.78110 Cer(d18:0/24:0) 0.00254 −27.27562 PC 18:0/20:4 0.00303 −16.02110 CE 20:3 0.00312 −19.02446 CE 17:1 0.00694 −16.90145 PC 18:0/20:3 0.00726 −17.20664 PC 18:0/18:1 0.00765 −18.18002 Cer(d18:0/22:0) 0.01158 −22.37263 PC 16:0/22:6 0.01180 −16.65050 LPC 18:1 0.01457 −14.45827 SM (d18:1/23:0) (d18:1/22:1-OH) 0.01920 −14.02360 CE 16:0 0.02427 −10.63490 Total CE 0.02745 −11.83333 SM (d18:1/24:0) (d18:1/23:1-OH) 0.02784 −13.83715 PC 18:1/18:2 0.03666 −10.83371 2b) Significant lipids in LURIC study sorted by p-value from subjects not undergoing statin treatment. Positive correlation Cer(d18:1/20:0) 0.00004 28.00357 Cer(d18:1/18:0) 0.00009 34.32550 LacCer(d18:1/20:0) 0.00010 32.91154 Cer(d18:1/24:1) 0.00039 23.37606 LacCer(d18:1/22:0) 0.00048 25.61851 LacCer(d18:1/18:0) 0.00144 29.19850 LacCer(d18:1/24:1) 0.00199 29.83279 PS O-18:2/16:0-alkenyl 0.00432 33.81177 Cer(d18:1/22:0) 0.00473 18.32126 PS O-16:0/18:2-alkenyl 0.00590 32.17190 PS O-16:1/18:2-alkyl 0.00590 32.17190 Total DAG 0.00794 31.67365 Cer(d18:1/16:0) 0.00952 16.71359 Total Cer 0.00932 15.44601 Total LacCer 0.01105 16.00541 LacCer(d18:1/24:0) 0.01989 19.85622 GlcCer(d18:1/20:0) 0.02288 17.74772 PC O-32:0 (KDdiA-PC) 0.02467 22.23265 GlcCer(d18:1/18:0) 0.02584 16.12961 GlcCer(d18:1/24:1) 0.03290 18.89331 GlcCer(d18:1/26:1) 0.04702 17.52675 Cer(d18:1/26:1) 0.04802 13.59618 Negative correlation Total PC 0.00921 −12.44220 CE 14:0 0.01090 −21.01258 CE 20:3 0.02157 −16.03606 CE 17:1 0.02204 −15.93952 PC 16:0/20:4 0.02256 −14.96966 PC 18:0/20:4 0.03376 −13.60917 Cer(d18:0/24:0) 0.03376 −21.87004 Total LPC 0.03443 −12.91576 PC 16:0/20:3 0.04337 −13.43056

TABLE 3 Table of significant lipid to lipid ratios in LURIC study sorted by p-value. Lipid names, p-values, % change both for positive and negative correlation are presented. Table 3a shows significant lipids from all study subjects. The significant lipids from subjects not undergoing statin treatment are listed in table 3b. Percentage Lipid name/lipid name p-value change 3a) Significant lipid to lipid ratios in LURIC study sorted by p-value from all study subjects. Positive correlation GlcCer(d18:1/26:0)/Total CE 0.0000000 12.8050228 GlcCer(d18:1/26:1)/Total CE 0.0000000 29.5576923 PS O-16:0/18:2-alkenyl/Total PS O 0.0000000 26.0723978 PS O-16:1/18:2-alkyl/Total PS O 0.0000000 26.0723978 PS O-18:2/16:0-alkenyl/Total PS O 0.0000000 28.7826715 Cer(d18:1/24:1)/Total PC 0.0000000 38.6612692 PC 16:0/16:0/Total PC 0.0000000 26.1328496 LacCer(d18:1/20:0)/Total PC 0.0000000 55.3076855 Cer(d18:1/24:1)/PC 16:0/20:4 0.0000000 52.6701270 Cer(d18:1/20:0)/Total PC 0.0000000 40.0940286 LacCer(d18:1/18:0)/Total PC 0.0000000 53.5855166 Cer(d18:1/18:0)/Total PC 0.0000000 48.2586905 Cer(d18:1/18:0)/PC 16:0/20:4 0.0000000 65.7060860 Cer(d18:1/24:1)/PC 16:0/18:1 0.0000000 34.2415459 PS O-18:0/18:2-alkenyl 0.0000000 18.7146206 (PS O-18:1/18:2-alkyl)/Total PS O Total LacCer/Total PC 0.0000001 40.0389043 Cer(d18:1/24:1)/PC 16:0/20:3 0.0000001 39.6033049 Cer(d18:1/22:0)/Total PC 0.0000001 31.3604924 Cer(d18:1/20:0)/PC 16:0/20:4 0.0000001 55.6619154 LacCer(d18:1/22:0)/Total PC 0.0000001 48.7385500 LacCer(d18:1/20:0)/PC 16:0/20:3 0.0000001 57.2475183 Cer(d18:1/18:0)/PC 16:0/20:3 0.0000002 53.4975004 LacCer(d18:1/20:0)/PC 18:0/20:3 0.0000002 56.3125345 LacCer(d18:1/18:0)/PC 18:0/20:3 0.0000002 58.4642532 Cer(d18:1/22:0)/PC 16:0/20:4 0.0000002 42.5755107 LacCer(d18:1/20:0)/PC 18:0/18:1 0.0000004 50.6042672 Cer(d18:1/24:1)/LPC 18:1 0.0000004 37.9852355 LacCer(d18:1/20:0)/PC 16:0/18:1 0.0000004 46.3528827 LacCer(d18:1/20:0)/PC 18:1/18:1 0.0000004 49.6378601 Cer(d18:1/18:0)/PC 16:0/18:1 0.0000004 42.1348344 Cer(d18:1/18:0)/PC 18:0/20:4 0.0000005 55.4207048 Cer(d18:1/24:1)/PC 18:0/20:4 0.0000005 40.7129933 Cer(d18:1/20:0)/PC 16:0/20:3 0.0000005 44.8451813 LacCer(d18:1/18:0)/PC 18:0/18:1 0.0000006 50.0860210 Cer(d18:1/20:0)/PC 16:0/18:1 0.0000006 35.2897482 LacCer(d18:1/20:0)/PC 18:1/18:2 0.0000007 47.4807020 Cer(d18:1/24:1)/Total LPC 0.0000007 37.5873387 Total Cer/Total PC 0.0000007 28.8638001 Cer(d18:1/20:0)/PC 18:0/20:4 0.0000007 46.2204972 LacCer(d18:1/18:0)/PC 18:1/18:1 0.0000007 47.3110220 LacCer(d18:1/18:0)/PC 16:0/20:3 0.0000007 55.5925567 LacCer(d18:1/24:1)/Total PC 0.0000008 56.5726916 Cer(d18:1/24:1)/PC 18:0/18:1 0.0000008 38.3563305 Cer(d18:1/24:1)/PC 18:0/20:3 0.0000013 39.6802706 LacCer(d18:1/22:0)/PC 16:0/20:4 0.0000013 57.8428560 LacCer(d18:1/18:0)/PC 16:0/18:1 0.0000014 45.2070855 LacCer(d18:1/22:0)/PC 18:0/20:3 0.0000014 50.7183795 Cer(d18:1/20:0)/PC 18:1/18:1 0.0000016 40.1833717 LacCer(d18:1/18:0)/PC 18:1/18:2 0.0000017 46.5169450 Cer(d18:1/22:0)/PC 16:0/20:3 0.0000018 33.7513783 Negative correlation Cer(d18:0/22:0)/Total CE 0.0000000 −11.7767698 Cer(d18:0/24:0)/Total CE 0.0000000 −18.9531828 Cer(d18:0/24:1)/Total CE 0.0000000 −1.2444181 CE 18:3/Cer(d18:1/24:1) 0.0000000 −35.9591235 Cer(d18:0/24:0)/Cer(d18:1/24:1) 0.0000004 −36.1288066 CE 18:3/Total Cer 0.0000021 −30.3105704 CE 18:3/PC 16:0/16:0 0.0000028 −30.4716021 CE 18:3/LacCer(d18:1/20:0) 0.0000028 −44.3063424 CE 20:3/Cer(d18:1/24:1) 0.0000034 −28.1216690 CE 18:3/PS O-16:0/18:2-alkenyl 0.0000036 −38.5094276 CE 18:3/PS O-16:1/18:2-alkyl 0.0000036 −38.5094276 CE 18:3/Total LacCer 0.0000037 −34.1308012 CE 18:3/Cer(d18:1/20:0) 0.0000037 −33.6203392 LPC 18:2/LacCer(d18:1/20:0) 0.0000040 −41.6604394 Cer(d18:0/24:0)/Total Cer 0.0000049 −31.4531758 Cer(d18:0/24:0)/Cer(d18:1/18:0) 0.0000052 −40.2960344 Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl 0.0000059 −40.8329837 Cer(d18:0/24:0)/PS O-16:1/18:2-alkyl 0.0000059 −40.8329837 Cer(d18:0/22:0)/Cer(d18:1/24:1) 0.0000064 −32.1886967 Cer(d18:0/24:0)/LacCer(d18:1/24:0) 0.0000067 −37.0287234 Cer(d18:0/22:0)/Cer(d18:1/18:0) 0.0000071 −36.5400374 CE 14:0/Cer(d18:1/24:1) 0.0000076 −29.9054883 CE 18:3/Cer(d18:1/16:0) 0.0000079 −32.5598715 CE 18:3/Cer(d18:1/18:0) 0.0000082 −38.1543108 Cer(d18:0/24:0)/Cer(d18:1/20:0) 0.0000087 −35.4631184 CE 18:3/GlcCer(d18:1/20:0) 0.0000087 −36.4665498 CE 18:3/Cer(d18:1/22:0) 0.0000101 −30.6595286 Cer(d18:0/24:0)/Cer(d18:1/22:0) 0.0000105 −32.4086711 Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl 0.0000114 −37.4226868 Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl 0.0000114 −37.4226868 CE 18:3/PC O-34:1 0.0000122 −33.4941413 CE 18:3/Total CE 0.0000151 −20.2882080 PC 16:0/20:3/PS O-16:0/18:2-alkenyl 0.0000154 −29.5280580 PC 16:0/20:3/PS O-16:1/18:2-alkyl 0.0000154 −29.5280580 PC 18:1/18:2/PS O-16:0/18:2-alkenyl 0.0000155 −23.7129392 PC 18:1/18:2/PS O-16:1/18:2-alkyl 0.0000155 −23.7129392 Cer(d18:0/22:0)/Cer(d18:1/20:0) 0.0000165 −31.5266149 CE 18:3/LacCer(d18:1/22:0) 0.0000174 −37.1525849 LPC 18:2/PS O-16:0/18:2-alkenyl 0.0000175 −30.3450779 LPC 18:2/PS O-16:1/18:2-alkyl 0.0000175 −30.3450779 CE 17:1/Cer(d18:1/24:1) 0.0000186 −24.8594478 CE 16:0/Cer(d18:1/24:1) 0.0000188 −19.2509390 CE 18:3/GlcCer(d18:1/18:0) 0.0000221 −35.5601491 CE 18:3/LacCer(d18:1/18:0) 0.0000326 −37.4416706 CE 18:3/PS O-16:0/18:1-alkenyl 0.0000326 −34.1672013 (PS O-16:1/18:1-alkyl) CE 18:3/Cer(d18:1/24:0) 0.0000331 −27.4645341 Cer(d18:0/22:0)/Cer(d18:1/22:0) 0.0000377 −28.0899033 LPC 16:0/LacCer(d18:1/20:0) 0.0000391 −35.6188222 Cer(d18:0/24:0)/Cer(d18:1/16:0) 0.0000446 −34.6695918 Total LPC/Total LacCer 0.0000469 −26.0881987 3b) Significant lipid to lipid ratios in LURIC study sorted by p-value from subjects not undergoing statin treatment Positive correlation GlcCer(d18:1/26:0)/Total CE 0.000000 13.329316 GlcCer(d18:1/26:1)/Total CE 0.000000 35.356592 PS O-16:0/18:2-alkenyl/Total PS O 0.000000 26.426208 PS O-16:1/18:2-alkyl/Total PS O 0.000000 26.426208 PS O-18:2/16:0-alkenyl/Total PS O 0.000000 29.590682 Cer(d18:1/24:1)/Total PC 0.000000 43.040800 Cer(d18:1/24:1)/PC 16:0/20:4 0.000000 59.876132 Cer(d18:1/24:1)/PC 18:0/20:4 0.000000 46.504629 Cer(d18:1/20:0)/Total PC 0.000000 46.216544 PC 16:0/16:0/Total PC 0.000000 28.139641 Total DAG/Total LPC 0.000000 57.426180 Cer(d18:1/18:0)/PC 16:0/20:4 0.000000 73.073704 Cer(d18:1/18:0)/Total PC 0.000000 53.769121 Cer(d18:1/22:0)/Total PC 0.000000 34.704834 Cer(d18:1/22:0)/PC 16:0/20:4 0.000000 47.860118 Cer(d18:1/20:0)/PC 16:0/20:4 0.000000 64.045788 LacCer(d18:1/20:0)/Total PC 0.000000 55.686130 Cer(d18:1/18:0)/PC 18:0/20:4 0.000000 61.943925 Cer(d18:1/20:0)/PC 18:0/20:4 0.000000 53.941480 Cer(d18:1/24:1)/PC 16:0/18:1 0.000000 35.646899 Cer(d18:1/24:1)/PC 16:0/20:3 0.000000 44.382440 Cer(d18:1/18:0)/PC 16:0/20:3 0.000001 61.224227 Cer(d18:1/20:0)/PC 16:0/20:3 0.000001 52.891307 Cer(d18:1/24:1)/PC 18:0/18:1 0.000001 42.115354 Cer(d18:1/22:0)/PC 18:0/20:4 0.000001 36.949978 Cer(d18:1/20:0)/PC 18:0/18:1 0.000004 47.396000 SM (d18:1/17:0) (d18:1/16:1-OH)/Total PC O 0.000004 39.660537 PC O-32:0 (KDdiA-PC)/Total PC O 0.000005 46.418572 Cer(d18:1/20:0)/PC 16:0/18:1 0.000005 37.704904 Cer(d18:1/24:1)/LPC 18:1 0.000005 40.092181 Cer(d18:1/24:1)/PC 18:1/18:1 0.000005 38.720001 Cer(d18:1/18:0)/PC 16:0/18:1 0.000006 44.307054 LacCer(d18:1/18:0)/Total PC 0.000006 50.539560 Cer(d18:1/20:0)/PC 18:1/18:1 0.000006 44.664936 Cer(d18:1/18:0)/PC 18:0/18:1 0.000006 54.563687 LacCer(d18:1/20:0)/PC 18:0/20:4 0.000007 66.072520 Cer(d18:1/24:1)/PC 18:0/20:3 0.000007 44.320399 LacCer(d18:1/20:0)/PC 16:0/20:3 0.000008 56.972652 Cer(d18:1/22:0)/PC 16:0/20:3 0.000008 37.291136 PC 16:0/16:0/PC 16:0/20:4 0.000009 42.512740 PS O-16:0/18:2-alkenyl/Total PC 0.000009 53.059092 PS O-16:1/18:2-alkyl/Total PC 0.000009 53.059092 LacCer(d18:1/20:0)/PC 18:0/20:3 0.000009 55.403274 Total Cer/Total PC 0.000011 29.949121 Total LacCer/Total PC 0.000012 37.815865 Total DAG/Total PC 0.000012 57.897424 Cer(d18:1/18:0)/PC 18:1/18:1 0.000012 49.427440 LacCer(d18:1/20:0)/PC 18:1/18:2 0.000013 48.432788 Total DAG/Total PC O 0.000014 52.258466 Cer(d18:1/24:1)/Total LPC 0.000014 38.684740 Negative correlation Cer(d18:0/22:0)/Total CE 0.000000 −6.628640 Cer(d18:0/24:0)/Total CE 0.000000 −13.307278 DAG 16:0/18:1/Total DAG 0.000000 −17.305276 CE 18:3/Cer(d18:1/24:1) 0.000000 −37.975733 CE 16:0/Cer(d18:1/24:1) 0.000001 −23.257135 Cer(d18:0/24:0)/Cer(d18:1/24:1) 0.000003 −36.760563 CE 20:4/Cer(d18:1/24:1) 0.000004 −28.636879 CE 20:3/Cer(d18:1/24:1) 0.000007 −30.787067 CE 14:0/Cer(d18:1/24:1) 0.000008 −32.409429 Cer(d18:0/22:0)/Cer(d18:1/24:1) 0.000011 −34.086320 CE 20:4/Cer(d18:1/18:0) 0.000017 −31.313949 CE 18:3/LacCer(d18:1/20:0) 0.000017 −45.652383 CE 18:3/Cer(d18:1/20:0) 0.000019 −34.938575 CE 17:1/Cer(d18:1/24:1) 0.000020 −27.626744 CE 18:3/Cer(d18:1/18:0) 0.000022 −39.797182 CE 18:3/PS O-16:0/18:2-alkenyl 0.000023 −40.724368 CE 18:3/PS O-16:1/18:2-alkyl 0.000023 −40.724368 CE 18:3/PC O-34:1 0.000025 −36.379680 CE 18:3/Total DAG 0.000025 −40.454303 CE 18:3/PC 16:0/16:0 0.000026 −31.169278 CE 18:2/Cer(d18:1/24:1) 0.000027 −23.500781 Cer(d18:0/22:0)/Cer(d18:1/18:0) 0.000031 −38.447200 CE 18:3/Total Cer 0.000034 −30.424215 Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl 0.000045 −37.930990 Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl 0.000045 −37.930990 LPC 18:2/LacCer(d18:1/20:0) 0.000045 −40.925010 PC 18:0/20:4/PS O-16:0/18:2-alkenyl 0.000046 −31.467609 PC 18:0/20:4/PS O-16:1/18:2-alkyl 0.000046 −31.467609 PC 16:0/20:4/Total DAG 0.000046 −36.637600 CE 16:0/Cer(d18:1/18:0) 0.000047 −26.423623 Cer(d18:0/22:0)/Cer(d18:1/20:0) 0.000051 −32.923442 PC 16:0/20:3/PS O-16:0/18:2-alkenyl 0.000051 −30.612669 PC 16:0/20:3/PS O-16:1/18:2-alkyl 0.000051 −30.612669 Cer(d18:0/24:0)/Cer(d18:1/18:0) 0.000052 −41.966868 CE 17:1/Cer(d18:1/18:0) 0.000062 −30.441023 CE 20:4/LacCer(d18:1/20:0) 0.000062 −34.937892 Cer(d18:0/24:0)/Cer(d18:1/20:0) 0.000077 −36.424116 CE 18:3/GlcCer(d18:1/20:0) 0.000078 −38.775015 PC 16:0/20:4/PS O-16:0/18:2-alkenyl 0.000079 −31.649115 PC 16:0/20:4/PS O-16:1/18:2-alkyl 0.000079 −31.649115 Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl 0.000082 −40.069396 Cer(d18:0/24:0)/PS O-16:1/18:2-alkyl 0.000082 −40.069396 Cer(d18:0/24:0)/Total Cer 0.000091 −30.775984 Cer(d18:0/22:0)/Total DAG 0.000096 −39.183597 CE 18:3/Cer(d18:1/16:0) 0.000101 −32.942871 PC 18:1/18:2/PS O-16:0/18:2-alkenyl 0.000106 −24.667433 PC 18:1/18:2/PS O-16:1/18:2-alkyl 0.000106 −24.667433 CE 18:3/Cer(d18:1/22:0) 0.000109 −30.876034 CE 14:0/Cer(d18:1/18:0) 0.000117 −35.001327 PC 18:0/20:3/PS O-16:0/18:2-alkenyl 0.000119 −29.825217

TABLE 4 Table of significant lipid to clinical ratios in LURIC study sorted by p-value. Lipid names and clinical measurement, p-values and percentage change both for positive and negative correlation are presented. Table 4a shows significant lipids from all study subjects. The significant lipids from subjects not undergoing statin treatment are listed in table 4b. Percentage Lipid name/clinical measurement p-value change 4a) Significant lipid to clinical ratios in LURIC study sorted by p-value from all study subjects. Positive correlation LacCer(d18:1/20:0)/apolipoprotein A-I 0.00000 43.04967 LacCer(d18:1/20:0)/total cholesterol 0.00000 41.46919 LacCer(d18:1/22:0)/total cholesterol 0.00000 32.24669 LacCer(d18:1/20:0)/triglycerides 0.00001 54.05564 LacCer(d18:1/22:0)/apolipoprotein A-I 0.00001 32.96039 Cer(d18:1/18:0)/triglycerides 0.00001 40.06643 Cer(d18:1/18:0)/total cholesterol 0.00002 31.56099 LacCer(d18:1/20:0)/apolipoprotein B 0.00002 37.03057 Cer(d18:1/20:0)/triglycerides 0.00003 34.46408 Cer(d18:1/20:0)/total cholesterol 0.00003 25.23403 LacCer(d18:1/18:0)/triglycerides 0.00004 52.60972 LacCer(d18:1/24:1)/triglycerides 0.00004 60.79934 LacCer(d18:1/22:0)/apolipoprotein B 0.00004 29.83861 Cer(d18:1/20:0)/apolipoprotein A-I 0.00004 31.35691 LacCer(d18:1/18:0)/total cholesterol 0.00005 38.87517 Cer(d18:1/24:1)/triglycerides 0.00005 32.27486 LacCer(d18:1/20:0)/HDL cholesterol 0.00006 39.32664 LacCer(d18:1/22:0)/triglycerides 0.00006 49.54525 LacCer(d18:1/18:0)/apolipoprotein A-I 0.00007 41.61371 LacCer(d18:1/24:1)/total cholesterol 0.00007 36.35465 Total LacCer/total cholesterol 0.00008 23.69633 Cer(d18:1/18:0)/apolipoprotein A-I 0.00008 37.38237 Cer(d18:1/24:1)/total cholesterol 0.00011 21.94547 Cer(d18:1/24:1)/apolipoprotein A-I 0.00011 27.12684 Cer(d18:1/18:0)/apolipoprotein B 0.00017 27.27914 LacCer(d18:1/20:0)/total-c/HDL-c 0.00018 33.48550 LacCer(d18:1/24:1)/apolipoprotein A-I 0.00018 36.33201 Total LacCer/triglycerides 0.00022 38.52220 LacCer(d18:1/24:1)/apolipoprotein B 0.00022 33.98045 LacCer(d18:1/18:0)/apolipoprotein B 0.00022 33.69302 LacCer(d18:1/22:0)/HDL cholesterol 0.00024 29.19905 Total LacCer/apolipoprotein A-I 0.00025 25.87122 PC O-32:0 (KDdiA-PC)/triglycerides 0.00029 48.31010 LacCer(d18:1/20:0)/LDL cholesterol 0.00031 31.95659 Cer(d18:1/22:0)/triglycerides 0.00037 28.23304 Total Cer/triglycerides 0.00043 26.80115 LacCer(d18:1/22:0)/LDL cholesterol 0.00050 24.24011 Cer(d18:1/20:0)/apolipoprotein B 0.00052 20.79615 LacCer(d18:1/22:0)/total-c/HDL-c 0.00061 27.84411 Total LacCer/apolipoprotein B 0.00065 20.63257 PC O-34:1/triglycerides 0.00066 38.80785 PC O-32:0 (KDdiA-PC)/apolipoprotein A-I 0.00066 36.57664 Cer(d18:1/20:0)/HDL cholesterol 0.00067 30.67863 LacCer(d18:1/18:0)/total-c/HDL-c 0.00073 30.67436 PS O-18:2/16:0-alkenyl/HDL cholesterol 0.00073 57.54661 LacCer(d18:1/18:0)/HDL cholesterol 0.00079 38.04132 Cer(d18:1/16:0)/triglycerides 0.00083 28.51323 Cer(d18:1/18:0)/HDL cholesterol 0.00085 36.16483 Cer(d18:1/18:0)/total-c/HDL-c 0.00087 21.75165 LacCer(d18:1/20:0)/apoA1/apoB 0.00090 32.36387 Negative correlation CE 18:3/apolipoprotein B 0.00016 −25.31907 CE 18:3/total cholesterol 0.00026 −23.67665 CE 18:3/LDL cholesterol 0.00052 −27.21759 CE 18:3/apolipoprotein A-I 0.00054 −23.09816 CE 18:3/LDL-c/HDL-c 0.00060 −31.33346 CE 18:3/apoA1/apoB 0.00068 −27.55825 CE 18:3/total-c/HDL-c 0.00070 −27.28733 CE 18:3/HDL cholesterol 0.00072 −24.04060 Cer(d18:0/24:0)/apolipoprotein B 0.00181 −26.25271 CE 14:0/LDL-c/HDL-c 0.00241 −27.07756 Cer(d18:0/24:0)/total cholesterol 0.00281 −24.73116 Cer(d18:0/24:0)/total-c/HDL-c 0.00347 −27.84469 CE 14:0/total-c/HDL-c 0.00356 −23.22809 PC 16:0/20:3/total-c/HDL-c 0.00364 −19.07121 CE 14:0/apolipoprotein B 0.00375 −19.76414 Total PC/LDL-c/HDL-c 0.00402 −21.24583 Cer(d18:0/24:0)/LDL-c/HDL-c 0.00502 −33.76759 Total PC/apolipoprotein B 0.00513 −12.91145 Total PC/total-c/HDL-c 0.00535 −16.13350 CE 20:3/LDL-c/HDL-c 0.00547 −21.16631 PC 16:0/20:4/apolipoprotein B 0.00576 −15.39247 PC 16:0/20:4/total-c/HDL-c 0.00682 −18.06323 Cer(d18:0/22:0)/LDL-c/HDL-c 0.00748 −29.05891 Total PC/total cholesterol 0.00760 −10.60231 Cer(d18:0/22:0)/total-c/HDL-c 0.00765 −23.72182 PC 16:0/20:3/LDL-c/HDL-c 0.00784 −22.56336 PC 18:0/20:3/total-c/HDL-c 0.00794 −17.83335 CE 14:0/total cholesterol 0.00816 −17.44260 CE 20:3/total-c/HDL-c 0.00819 −17.34055 PC 16:0/20:4/LDL-c/HDL-c 0.00828 −21.49898 CE 17:1/LDL-c/HDL-c 0.00832 −21.33179 CE 14:0/LDL cholesterol 0.00832 −20.55966 Cer(d18:0/24:0)/LDL cholesterol 0.00848 −30.08563 LPC 18:2/LDL-c/HDL-c 0.00873 −21.70580 Total LPC/LDL-c/HDL-c 0.00881 −20.39287 PC 18:0/20:3/LDL-c/HDL-c 0.00947 −22.19538 Cer(d18:0/24:0)/apolipoprotein A-I 0.01052 −22.39254 Cer(d18:0/22:0)/apolipoprotein B 0.01094 −21.02661 CE 20:3/apolipoprotein B 0.01105 −15.18746 PC 16:0/20:4/total cholesterol 0.01131 −13.37554 Total LPC/total-c/HDL-c 0.01183 −16.82334 LPC 18:2/apoA1/apoB 0.01189 −19.23316 PC 16:0/20:3/apolipoprotein B 0.01273 −15.11848 CE 16:1/apolipoprotein B 0.01456 −26.99397 Cer(d18:0/24:0)/HDL cholesterol 0.01507 −23.10073 CE 16:1/LDL cholesterol 0.01581 −29.68280 PC 18:0/18:1/LDL-c/HDL-c 0.01581 −25.72002 CE 20:5/HDL cholesterol 0.01584 −23.45029 CE 20:5/apolipoprotein B 0.01619 −27.03163 LPC 18:2/LDL cholesterol 0.01648 −17.26480 4b) Significant lipid to clinical ratios in LURIC study sorted by p-value from subjects not undergoing statin treatment. Positive correlation Cer(d18:1/20:0)/total cholesterol 0.00000 38.78716 Cer(d18:1/20:0)/apolipoprotein B 0.00000 35.29473 Cer(d18:1/18:0)/total cholesterol 0.00000 44.87416 Cer(d18:1/24:1)/total cholesterol 0.00000 31.81230 Cer(d18:1/18:0)/apolipoprotein B 0.00000 41.46939 Cer(d18:1/20:0)/apolipoprotein A-I 0.00000 43.22923 Cer(d18:1/24:1)/apolipoprotein A-I 0.00000 36.15491 LacCer(d18:1/20:0)/apolipoprotein A-I 0.00001 47.52465 Cer(d18:1/18:0)/apolipoprotein A-I 0.00001 49.43693 LacCer(d18:1/20:0)/total cholesterol 0.00001 47.43498 Cer(d18:1/22:0)/total cholesterol 0.00001 26.21569 LacCer(d18:1/22:0)/total cholesterol 0.00001 36.57029 Cer(d18:1/24:1)/apolipoprotein B 0.00001 29.03655 LacCer(d18:1/22:0)/apolipoprotein A-I 0.00001 35.92744 Cer(d18:1/18:0)/triglycerides 0.00002 45.88769 Cer(d18:1/20:0)/triglycerides 0.00002 41.38147 LacCer(d18:1/20:0)/apolipoprotein B 0.00002 43.58427 Total Cer/total cholesterol 0.00002 22.69184 Total DAG/triglycerides 0.00003 42.61436 LacCer(d18:1/20:0)/LDL cholesterol 0.00003 42.53680 Cer(d18:1/18:0)/total-c/HDL-c 0.00003 32.87649 Cer(d18:1/24:1)/triglycerides 0.00004 37.38060 Cer(d18:1/20:0)/HDL cholesterol 0.00004 44.64484 LacCer(d18:1/22:0)/apolipoprotein B 0.00004 35.19942 Cer(d18:1/22:0)/apolipoprotein B 0.00005 24.23877 LacCer(d18:1/24:1)/total cholesterol 0.00006 43.51653 LacCer(d18:1/20:0)/HDL cholesterol 0.00006 45.00931 Total LacCer/total cholesterol 0.00007 27.38505 Cer(d18:1/18:0)/HDL cholesterol 0.00008 50.71310 Cer(d18:1/24:1)/HDL cholesterol 0.00008 36.69128 LacCer(d18:1/20:0)/triglycerides 0.00009 53.20068 LacCer(d18:1/24:1)/apolipoprotein B 0.00009 42.74199 Cer(d18:1/20:0)/total-c/HDL-c 0.00010 26.38965 LacCer(d18:1/22:0)/LDL cholesterol 0.00010 31.99739 Cer(d18:1/22:0)/apolipoprotein A-I 0.00011 28.85911 LacCer(d18:1/24:1)/triglycerides 0.00013 64.74061 Total Cer/apolipoprotein B 0.00015 21.06506 Total Cer/apolipoprotein A-I 0.00016 25.36811 Cer(d18:1/16:0)/total cholesterol 0.00016 27.48182 LacCer(d18:1/24:1)/apolipoprotein A-I 0.00017 41.35186 Cer(d18:1/20:0)/LDL cholesterol 0.00018 35.64759 LacCer(d18:1/18:0)/total cholesterol 0.00019 42.30922 LacCer(d18:1/22:0)/HDL cholesterol 0.00020 33.63345 LacCer(d18:1/18:0)/apolipoprotein A-I 0.00022 44.38352 Cer(d18:1/18:0)/LDL cholesterol 0.00023 42.34532 Total LacCer/apolipoprotein A-I 0.00031 28.05057 Total LacCer/apolipoprotein B 0.00035 24.82894 Cer(d18:1/22:0)/triglycerides 0.00039 32.11726 LacCer(d18:1/20:0)/total-c/HDL-c 0.00040 37.19117 Cer(d18:1/16:0)/apolipoprotein A-I 0.00042 30.73655 Negative correlation CE 18:3/apoA1/apoB 0.00213 −27.06449 CE 18:3/apolipoprotein A-I 0.00385 −21.63982 CE 18:3/apolipoprotein B 0.00576 −21.66212 CE 18:3/total cholesterol 0.00593 −20.58905 CE 18:3/HDL cholesterol 0.00621 −21.44255 CE 18:3/total-c/HDL-c 0.01166 −23.94433 CE 18:3/LDL-c/HDL-c 0.01395 −25.96634 CE 18:3/LDL cholesterol 0.01410 −22.13225 CE 14:0/total-c/HDL-c 0.02379 −20.97683 CE 14:0/LDL-c/HDL-c 0.03038 −22.86773 Cer(d18:0/24:0)/apolipoprotein B 0.03428 −21.10483 CE 14:0/apolipoprotein B 0.03640 −16.53355 Cer(d18:0/24:0)/total-c/HDL-c 0.04122 −23.22174 Cer(d18:0/24:0)/total cholesterol 0.04309 −19.71324 CE 14:0/apoA1/apoB 0.04354 −18.62921 CE 16:1/apoA1/apoB 0.04511 −23.73367

The biomarker ability of measured lipids was assessed also by calculating the sensitivity and specificity values for each lipid and their ratios to other lipids or classical biomarkers such as LDL-C and apolipoproteins. This ROC curve analysis revealed a number of biomarker candidates that have equal of higher than 60% sensitivity and specificity for predicting CVD complications (Tables 7-9).

TABLE 5 Significant lipids in LURIC study sorted by top sensitivity and specificity. Table 5a shows significant lipids from all study subjects. The significant lipids from subjects not undergoing statin treatment are listed in table 5b. Lipid name Sensitivity Specificity Percentage change 5a) Significant lipids in LURIC study sorted by top sensitivity and specificity from all study subjects. Positive correlation LacCer(d18:1/20:0) 66.03774 63.70370 29.52421 Total LacCer 63.15789 60.12270 13.83938 Cer(d18:1/24:1) 61.40351 60.12270 14.88898 Negative correlation Total PC 71.92982 63.29114 −16.07367 PC 16:0/20:3 71.42857 60.75949 −18.78110 Total LPC 70.17544 60.75949 −17.02070 PC 18:0/20:3 66.03774 63.05732 −17.20664 LPC 18:1 64.91228 60.12658 −14.45827 SM (d18:1/14:0) 63.79310 60.50955 −10.49341 (d18:1/13:1-OH) Cer(d18:0/22:0) 61.40351 61.34969 −22.37263 PC 16:0/18:2 61.40351 60.12658 −8.30551 PC 16:0/22:6 60.37736 61.14650 −16.65050 5b) Significant lipids in LURIC study sorted by top sensitivity and specificity from subjects not undergoing statin treatment. Positive correlation LacCer(d18:1/20:0) 70.45455 60.00000 29.52421 Cer(d18:1/24:1) 69.56522 60.34483 14.88898 Cer(d18:1/20:0) 65.21739 64.65517 17.32385 Cer(d18:1/22:0) 60.86957 62.93103 10.65433 GlcCer(d18:1/24:1) 60.86957 62.06897 12.75066 LacCer(d18:1/22:0) 60.00000 61.76471 22.75541 Negative correlation Total PC 69.56522 61.60714 −16.07367 PC 16:0/20:3 64.44444 60.71429 −18.78110 SM (d18:1/14:0) 63.82979 60.71429 −10.49341 (d18:1/13:1-OH) LPC 18:1 63.04348 60.71429 −14.45827 Total LPC 63.04348 60.71429 −17.02070 PC O-40:3 60.97561 60.43956 −1.88354 PC 16:0/20:4 60.86957 60.71429 −18.06547

TABLE 6 Table of significant lipid to lipid ratios in LURIC study sorted by top sensitivity and specificity. Table 6a shows significant lipids from all study subjects. The significant lipids from subjects not undergoing statin treatment are listed in table 6b. Lipid name/Lipid name Sensitivity Specificity Percentage change 6a) Table of significant lipid to lipid ratios in LURIC study sorted by top sensitivity and specificity from all study subjects. Positive correlation PS O-16:0/18:2-alkenyl/Total PS O 98.24561 63.92405 26.07240 PS O-16:1/18:2-alkyl/Total PS O 98.24561 63.92405 26.07240 PS O-18:2/16:0-alkenyl/Total PS O 85.45455 70.66667 28.78267 LacCer(d18:1/20:0)/PC 16:0/20:3 80.39216 65.11628 57.24752 CE 18:2/CE 18:3 79.31034 60.37736 26.34366 LacCer(d18:1/20:0)/Total LPC 78.84615 61.24031 53.68111 LacCer(d18:1/20:0)/Total PC 78.84615 64.34109 55.30769 LacCer(d18:1/20:0)/PC 18:0/20:3 77.08333 60.15625 56.31253 Cer(d18:1/24:1)/LPC 18:2 75.92593 63.69427 48.79017 CE 16:0/CE 18:3 75.86207 60.37736 25.64615 LacCer(d18:1/20:0)/Total SM 75.47170 60.60606 29.12486 Cer(d18:1/16:0)/Total PC 75.00000 61.14650 32.91697 Cer(d18:1/18:0)/PC 16:0/20:4 75.00000 63.05732 65.70609 Cer(d18:1/18:0)/Total LPC 75.00000 61.14650 45.10211 Cer(d18:1/24:1)/SM (d18:1/24:0) (d18:1/23:1- 75.00000 62.42038 36.77357 OH) Cer(d18:1/24:1)/Total LPC 75.00000 63.05732 37.58734 Cer(d18:1/24:1)/Total PC 75.00000 60.50955 38.66127 LacCer(d18:1/18:0)/Total LPC 75.00000 60.13072 55.13308 LacCer(d18:1/20:0)/SM (d18:1/17:1-OH) 75.00000 62.60163 36.30844 LacCer(d18:1/20:0)/SM (d18:1/18:0) 75.00000 62.60163 36.30844 Total Cer/Total PC 75.00000 63.92405 28.86380 LacCer(d18:1/24:0)/Total LPC 74.54545 61.94030 36.27249 LacCer(d18:1/20:0)/PC 18:2/18:2 74.46809 60.31746 47.81762 PC 16:0/16:0/Total PC 73.68421 65.18987 26.13285 Cer(d18:1/22:0)/Total PC 73.21429 61.78344 31.36049 GlcCer(d18:1/20:0)/Total PC 73.21429 60.50955 36.81380 LacCer(d18:1/16:0)/Total LPC 73.21429 60.50955 26.35547 LacCer(d18:1/18:0)/Total PC 73.21429 61.43791 53.58552 LacCer(d18:1/22:0)/SM (d18:1/14:0) 73.21429 60.00000 30.07361 (d18:1/13:1-OH) Total LacCer/Total PC 73.21429 60.50955 40.03890 LacCer(d18:1/20:0)/PC 16:0/18:1 73.07692 62.01550 46.35288 LacCer(d18:1/20:0)/PC 18:1/18:2 73.07692 60.46512 47.48070 Cer(d18:1/26:0)/PC O-40:0 72.72727 62.22222 9.97951 LacCer(d18:1/22:0)/Total LPC 72.72727 64.92537 41.61463 LacCer(d18:1/24:1)/Total LPC 72.72727 60.44776 45.29327 PC O-18:0/18:2-alkyl/PC O-36:5 72.72727 61.18421 25.99402 PC O-32:0 (KDdiA-PC)/PC O-38:5 72.54902 61.36364 34.00112 Cer(d18:1/22:0)/LPC 18:2 72.22222 61.78344 41.72749 LacCer(d18:1/22:0)/PC 16:0/20:3 72.22222 66.41791 51.38070 LacCer(d18:1/24:0)/PC 16:0/20:3 72.22222 61.19403 39.48118 Cer(d18:1/24:1)/Total CE 71.92982 61.39241 31.32927 PC 16:0/16:0/PC 16:0/20:4 71.92982 61.39241 38.61464 PC 16:0/18:2/Total PC 71.92982 60.75949 8.88941 Total LacCer/Total PC O 71.92982 60.24845 17.49430 Cer(d18:1/16:0)/LPC 18:1 71.42857 63.69427 30.55279 Cer(d18:1/18:0)/LPC 16:0 71.42857 61.14650 40.41256 Cer(d18:1/18:0)/LPC 18:1 71.42857 61.78344 45.33865 GlcCer(d18:1/20:0)/PC 16:0/20:4 71.42857 60.50955 46.04915 LacCer(d18:1/24:1)/Total PC O 71.42857 60.14493 21.09716 CE 19:1/Cer(d18:0/22:0) 71.15385 60.14493 39.21301 Negative correlation CE 18:3/LacCer(d18:1/20:0) 84.90566 62.87879 −44.30634 CE 18:3/Cer(d18:1/24:1) 80.70175 60.12658 −35.95912 Cer(d18:0/24:0)/Total Cer 80.70175 60.12270 −31.45318 LPC 18:2/LacCer(d18:1/20:0) 80.00000 64.34109 −41.66044 CE 18:3/Total CE 79.31034 61.00629 −20.28821 GlcCer(d18:1/26:0)/LacCer(d18:1/20:0) 79.16667 61.40351 −23.71130 CE 16:1/Cer(d18:1/20:0) 78.94737 61.39241 −33.84979 CE 16:1/Cer(d18:1/24:1) 77.19298 61.39241 −36.17234 CE 16:1/LacCer(d18:1/18:0) 77.19298 60.38961 −37.93756 CE 18:3/Cer(d18:1/20:0) 77.19298 60.75949 −33.62034 LPC 18:1/LacCer(d18:1/20:0) 76.92308 63.56589 −34.40451 CE 18:3/LacCer(d18:1/22:0) 76.78571 61.02941 −37.15258 CE 16:1/CE 19:1 75.47170 60.86957 −66.77151 CE 16:1/Cer(d18:1/18:0) 75.43860 61.39241 −39.42716 CE 16:1/LacCer(d18:1/16:0) 75.43860 60.12658 −30.53235 CE 16:1/Total LacCer 75.43860 60.75949 −34.87060 CE 18:3/PC 16:0/16:0 75.43860 60.00000 −30.47160 CE 18:3/PS O-16:0/18:2-alkenyl 75.43860 60.64516 −38.50943 CE 18:3/PS O-16:1/18:2-alkyl 75.43860 60.64516 −38.50943 CE 18:3/Total LacCer 75.43860 63.29114 −34.13080 Cer(d18:0/24:0)/Cer(d18:1/22:0) 75.43860 66.25767 −32.40867 CE 16:1/LacCer(d18:1/22:0) 75.00000 61.02941 −37.29505 CE 16:1/LacCer(d18:1/24:0) 75.00000 60.29412 −36.84318 GlcCer(d18:1/26:0)/LacCer(d18:1/22:0) 75.00000 60.86957 −20.64622 LPC 16:0/LacCer(d18:1/20:0) 75.00000 61.24031 −35.61882 Total LPC/Total LacCer 75.00000 61.78344 −26.08820 LPC 16:0/LacCer(d18:1/24:1) 74.54545 60.44776 −24.61533 CE 17:1/GlcCer(d18:1/24:1) 74.07407 60.25641 −24.94421 CE 16:1/GlcCer(d18:1/18:0) 73.68421 60.12658 −36.48698 CE 16:1/GlcCer(d18:1/20:0) 73.68421 60.75949 −38.30453 CE 16:1/PC 16:0/16:0 73.68421 60.64516 −29.10430 CE 18:1/Total LacCer 73.68421 60.12658 −20.87773 CE 18:3/Cer(d18:1/16:0) 73.68421 67.08861 −32.55987 CE 18:3/Cer(d18:1/22:0) 73.68421 61.39241 −30.65953 CE 20:3/Cer(d18:1/24:1) 73.68421 61.39241 −28.12167 Cer(d18:0/22:0)/Cer(d18:1/24:1) 73.68421 61.96319 −32.18870 Cer(d18:0/24:0)/Cer(d18:1/18:0) 73.68421 63.80368 −40.29603 Cer(d18:0/24:0)/Cer(d18:1/24:1) 73.68421 65.64417 −36.12881 Cer(d18:0/24:0)/GlcCer(d18:1/20:0) 73.68421 60.73620 −40.34621 CE 16:1/LacCer(d18:1/20:0) 73.58491 62.12121 −39.47654 CE 20:3/LacCer(d18:1/20:0) 73.58491 62.12121 −35.94789 CE 18:3/LacCer(d18:1/24:0) 73.21429 62.50000 −35.91095 CE 18:3/PS O-16:0/18:1-alkenyl (PS O- 73.21429 60.64516 −34.16720 16:1/18:1-alkyl) Cer(d18:0/22:0)/LacCer(d18:1/24:0) 73.21429 60.00000 −31.24173 Cer(d18:0/24:0)/LacCer(d18:1/24:0) 73.21429 62.85714 −37.02872 LPC 16:0/Total LacCer 73.21429 61.78344 −24.37341 SM (d18:1/24:0) (d18:1/23:1-OH)/Total Cer 73.21429 61.14650 −13.97590 Cer(d18:0/24:0)/SM (d18:1/17:0) (d18:1/16:1- 72.91667 60.00000 −44.87505 OH) LPC 16:0/LacCer(d18:1/22:0) 72.72727 67.16418 −27.47521 CE 17:1/LacCer(d18:1/18:0) 72.22222 60.52632 −27.90709 6b) Table of significant lipid to lipid ratios in LURIC study sorted by top sensitivity and specificity from subjects not undergoing statin treatment. Positive correlation PS O-16:0/18:2-alkenyl/Total PS O 97.82609 62.50000 26.42621 PS O-16:1/18:2-alkyl/Total PS O 97.82609 62.50000 26.42621 PS O-18:2/16:0-alkenyl/Total PS O 88.63636 72.64151 29.59068 Cer(d18:1/24:1)/LPC 18:2 81.39535 62.16216 51.68188 CE 18:2/CE 18:3 80.85106 60.52632 24.84398 Cer(d18:1/18:0)/PC 16:0/20:4 80.00000 62.16216 73.07370 Cer(d18:1/24:1)/PC 16:0/18:2 80.00000 60.36036 27.45405 Cer(d18:1/24:1)/Total LPC 80.00000 62.16216 38.68474 Cer(d18:1/24:1)/Total PC 80.00000 62.16216 43.04080 LacCer(d18:1/20:0)/PC 16:0/20:4 79.06977 68.42105 82.44735 LacCer(d18:1/20:0)/PC 16:0/20:3 78.57143 64.21053 56.97265 Cer(d18:1/16:0)/Total PC 77.77778 60.36036 34.07762 Cer(d18:1/18:0)/LPC 18:1 77.77778 60.36036 45.52166 Cer(d18:1/24:1)/SM (d18:1/24:0) (d18:1/23:1-OH) 77.77778 60.17699 32.27927 Cer(d18:1/24:1)/PC O-40:3 77.50000 60.43956 20.46959 LacCer(d18:1/20:0)/Total LPC 76.74419 60.00000 49.56627 LacCer(d18:1/20:0)/Total PC 76.74419 63.15789 55.68613 Cer(d18:1/18:0)/Total CE 76.08696 60.17699 49.43112 Cer(d18:1/20:0)/SM (d18:1/14:0) (d18:1/13:1-OH) 76.08696 60.71429 28.98221 Cer(d18:1/20:0)/Total PC O 76.08696 60.86957 29.01076 Cer(d18:1/24:1)/Total CE 76.08696 60.17699 39.34990 PS O-16:0/18:2-alkenyl/Total PC O 76.08696 61.60714 20.69456 PS O-16:1/18:2-alkyl/Total PC O 76.08696 61.60714 20.69456 Cer(d18:1/16:0)/LPC 18:1 75.55556 60.36036 26.36925 Cer(d18:1/18:0)/Total LPC 75.55556 63.06306 44.40414 Cer(d18:1/22:0)/Total PC 75.55556 63.06306 34.70483 Total Cer/Total PC 75.55556 61.60714 29.94912 LacCer(d18:1/20:0)/Total SM 75.00000 61.61616 27.48798 LacCer(d18:1/24:0)/Total LPC 75.00000 67.01031 34.46432 PS O-18:2/16:0-alkenyl/Total PC O 75.00000 60.37736 21.65448 CE 16:0/CE 18:3 74.46809 61.40351 25.51627 CE 18:0/CE 18:3 74.46809 65.13761 29.77017 Cer(d18:1/22:0)/LPC 18:2 74.41860 60.36036 44.40272 LacCer(d18:1/20:0)/PC 18:1/18:2 74.41860 64.21053 48.43279 LacCer(d18:1/20:0)/SM (d18:1/17:1-OH) 74.41860 60.86957 34.96305 LacCer(d18:1/20:0)/SM (d18:1/18:0) 74.41860 60.86957 34.96305 LacCer(d18:1/20:0)/PC 18:0/20:3 74.35897 62.76596 55.40327 Cer(d18:1/18:0)/SM (d18:1/14:0) (d18:1/13:1-OH) 73.91304 74.10714 34.95249 Cer(d18:1/18:0)/SM (d18:1/17:2-OH) 73.91304 60.00000 25.13954 Cer(d18:1/18:0)/SM (d18:1/18:1) 73.91304 60.00000 25.13954 Cer(d18:1/24:1)/SM (d18:1/23:0) (d18:1/22:1-OH) 73.91304 60.71429 29.95139 PC 16:0/16:0/PC 16:0/20:4 73.91304 60.71429 42.51274 Cer(d18:1/20:0)/Total LPC 73.33333 61.26126 36.63159 Cer(d18:1/24:1)/PC 16:0/20:4 73.33333 62.16216 59.87613 Cer(d18:1/24:1)/PC 18:0/18:2 73.33333 62.16216 29.37710 Cer(d18:1/24:1)/PC 18:1/18:2 73.33333 61.26126 36.88389 Cer(d18:1/24:1)/SM (d18:1/17:1-OH) 73.33333 64.42308 23.41916 Cer(d18:1/24:1)/SM (d18:1/18:0) 73.33333 64.42308 23.41916 LacCer(d18:1/20:0)/PC 18:0/18:1 73.17073 62.10526 50.18761 LacCer(d18:1/20:0)/Total CE 72.72727 62.62626 53.71776 Negative correlation DAG 16:0/18:1/Total DAG 88.88889 61.16505 −17.30528 CE 18:3/LacCer(d18:1/20:0) 86.36364 66.66667 −45.65238 LPC 18:2/LacCer(d18:1/20:0) 82.92683 62.10526 −40.92501 CE 18:3/Cer(d18:1/24:1) 82.60870 61.06195 −37.97573 GlcCer(d18:1/26:0)/LacCer(d18:1/20:0) 80.48780 64.77273 −26.83109 CE 14:0/Total DAG 80.00000 61.16505 −39.38279 CE 18:3/Total CE 78.72340 62.28070 −19.58872 PC 18:0/20:3/PS O-16:0/18:2-alkenyl 78.57143 60.36036 −29.82522 PC 18:0/20:3/PS O-16:1/18:2-alkyl 78.57143 60.36036 −29.82522 CE 16:1/Cer(d18:1/24:1) 78.26087 61.06195 −37.53854 CE 18:3/PS O-16:0/18:2-alkenyl 78.26087 60.90909 −40.72437 CE 18:3/PS O-16:1/18:2-alkyl 78.26087 60.90909 −40.72437 CE 18:3/Total LacCer 78.26087 60.17699 −32.64187 Cer(d18:0/24:0)/Total Cer 78.26087 62.06897 −30.77598 GlcCer(d18:1/26:0)/LacCer(d18:1/22:0) 78.04878 61.79775 −21.27116 CE 18:3/LacCer(d18:1/22:0) 77.77778 63.63636 −34.76187 CE 20:3/LacCer(d18:1/20:0) 77.27273 60.60606 −36.58675 PC O-40:3/PS O-18:2/16:0-alkenyl 76.92308 62.79070 −28.59638 LPC 18:1/LacCer(d18:1/20:0) 76.74419 60.00000 −33.84809 CE 16:0/Cer(d18:1/24:1) 76.08696 61.06195 −23.25713 CE 16:1/Cer(d18:1/18:0) 76.08696 63.71681 −41.11023 CE 16:1/Cer(d18:1/20:0) 76.08696 66.37168 −34.98271 CE 16:1/GlcCer(d18:1/24:1) 76.08696 60.17699 −39.81218 CE 16:1/LacCer(d18:1/18:0) 76.08696 60.36036 −37.28535 CE 18:3/Cer(d18:1/20:0) 76.08696 63.71681 −34.93858 CE 18:3/Cer(d18:1/22:0) 76.08696 61.94690 −30.87603 Cer(d18:0/22:0)/Cer(d18:1/24:1) 76.08696 61.20690 −34.08632 Cer(d18:0/22:0)/Total GlcCer 76.08696 60.34483 −28.95910 Cer(d18:0/24:0)/Cer(d18:1/20:0) 76.08696 60.34483 −36.42412 PC 18:0/20:3/PS O-18:2/16:0-alkenyl 75.60976 60.95238 −29.83126 CE 16:1/LacCer(d18:1/24:0) 75.55556 60.60606 −36.83943 CE 18:3/LacCer(d18:1/24:0) 75.55556 60.60606 −37.84417 PC 18:1/18:2/Total Cer 75.55556 60.71429 −19.58862 SM (d18:1/23:0) (d18:1/22:1-OH)/Total DAG 75.55556 63.10680 −37.45206 SM (d18:1/24:0) (d18:1/23:1-OH)/Total Cer 75.55556 61.94690 −11.07113 Total CE/Total DAG 75.55556 66.01942 −31.35698 Total LPC/Total LacCer 75.55556 69.36937 −23.44706 CE 16:1/CE 19:1 75.00000 60.82474 −56.18741 CE 20:5/LacCer(d18:1/20:0) 75.00000 61.61616 −37.54401 LPC 16:0/LacCer(d18:1/24:0) 75.00000 60.82474 −25.29458 CE 15:0/Cer(d18:1/20:0) 74.41860 60.57692 −21.94830 CE 16:0/Cer(d18:1/18:0) 73.91304 61.06195 −26.42362 CE 16:1/Total LacCer 73.91304 63.71681 −33.50629 CE 18:2/Cer(d18:1/20:0) 73.91304 61.94690 −19.85408 CE 18:3/Cer(d18:1/24:0) 73.91304 61.40351 −28.00256 CE 18:3/GlcCer(d18:1/20:0) 73.91304 65.48673 −38.77502 CE 18:3/PC 16:0/16:0 73.91304 63.63636 −31.16928 CE 20:3/Cer(d18:1/24:1) 73.91304 61.06195 −30.78707 CE 20:4/GlcCer(d18:1/20:0) 73.91304 62.83186 −30.95570 CE 20:4/GlcCer(d18:1/24:1) 73.91304 61.06195 −28.82844

TABLE 7 Table of significant lipid to clinical ratios in LURIC study sorted by top sensitivity and specificity. Table 7a shows significant lipids from all study subjects. The significant lipids from subjects not undergoing statin treatment are listed in table 7b. Percentage Lipid name/Clinical measurement Sensitivity Specificity change 7a) Table of significant lipid to clinical ratios in LURIC study sorted by top sensitivity and specificity from all study subjects. Positive correlation LacCer(d18:1/20:0)/ 75.47170 61.48148 43.04967 apolipoprotein A-I Cer(d18:1/24:1)/total cholesterol 70.17544 60.73620 21.94547 Cer(d18:1/24:1)/triglycerides 70.17544 60.12270 32.27486 LacCer(d18:1/20:0)/HDL cholesterol 69.81132 62.22222 39.32664 LacCer(d18:1/20:0)/apoA1/apoB 67.92453 61.48148 32.36387 LacCer(d18:1/22:0)/HDL cholesterol 67.85714 60.00000 29.19905 Cer(d18:1/16:0)/triglycerides 66.66667 61.34969 28.51323 Cer(d18:1/22:0)/apolipoprotein B 66.66667 60.12270 14.20773 Cer(d18:1/24:1)/apolipoprotein B 66.66667 62.57669 18.05222 Total Cer/total cholesterol 66.66667 65.85366 15.12131 LacCer(d18:1/20:0)/LDL-c/HDL-c 66.03774 60.74074 23.54484 LacCer(d18:1/20:0)/total cholesterol 66.03774 62.96296 41.46919 LacCer(d18:1/20:0)/total-c/HDL-c 66.03774 60.00000 33.48550 Cer(d18:1/18:0)/triglycerides 64.91228 60.73620 40.06643 Cer(d18:1/22:0)/total cholesterol 64.91228 61.34969 17.20335 Cer(d18:1/24:1)/total-c/HDL-c 64.91228 61.34969 13.34657 GlcCer(d18:1/24:0)/total cholesterol 64.91228 61.96319 17.04608 LacCer(d18:1/18:0)/ 64.91228 60.37736 41.61371 apolipoprotein A-I LacCer(d18:1/18:0)/total cholesterol 64.91228 61.00629 38.87517 Total LacCer/total cholesterol 64.91228 65.03067 23.69633 LacCer(d18:1/22:0)/apoA1/apoB 64.28571 62.85714 24.17423 LacCer(d18:1/22:0)/ 64.28571 62.85714 32.96039 apolipoprotein A-I LacCer(d18:1/22:0)/total cholesterol 64.28571 60.00000 32.24669 LacCer(d18:1/24:0)/apoA1/apoB 64.28571 62.14286 15.23336 LacCer(d18:1/20:0)/LDL cholesterol 64.15094 61.48148 31.95659 LacCer(d18:1/20:0)/apolipoprotein B 64.15094 60.00000 37.03057 PS O-18:2/16:0-alkenyl/triglycerides 63.63636 63.33333 99.88127 Cer(d18:1/20:0)/apolipoprotein B 63.15789 60.12270 20.79615 Cer(d18:1/20:0)/total cholesterol 63.15789 61.96319 25.23403 Cer(d18:1/22:0)/LDL cholesterol 63.15789 60.12270 11.30545 Cer(d18:1/24:1)/LDL cholesterol 63.15789 60.12270 14.20759 PS O-16:0/18:2-alkenyl/triglycerides 63.15789 63.29114 97.78843 PS O-16:1/18:2-alkyl/triglycerides 63.15789 63.29114 97.78843 Total GlcCer/total cholesterol 63.15789 60.73620 17.56924 Total LacCer/apolipoprotein A-I 63.15789 61.34969 25.87122 Total LacCer/apolipoprotein B 63.15789 61.96319 20.63257 LacCer(d18:1/24:1)/triglycerides 62.50000 62.14286 60.79934 PS O-16:0/18:1-alkenyl (PS 62.50000 61.39241 65.88734 O-16:1/18:1-alkyl)/triglycerides Cer(d18:1/20:0)/triglycerides 61.40351 60.73620 34.46408 Cer(d18:1/22:0)/triglycerides 61.40351 63.19018 28.23304 Cer(d18:1/24:1)/apolipoprotein A-I 61.40351 65.03067 27.12684 LacCer(d18:1/16:0)/triglycerides 61.40351 64.41718 25.01312 LacCer(d18:1/18:0)/HDL cholesterol 61.40351 60.37736 38.04132 LacCer(d18:1/18:0)/LDL cholesterol 61.40351 64.15094 28.50180 LacCer(d18:1/18:0)/apolipoprotein B 61.40351 61.00629 33.69302 LacCer(d18:1/18:0)/triglycerides 61.40351 62.26415 52.60972 Total Cer/apolipoprotein A-I 61.40351 63.41463 19.09944 Total GlcCer/apolipoprotein B 61.40351 60.12270 14.92180 Total LacCer/triglycerides 61.40351 62.57669 38.52220 LacCer(d18:1/22:0)/apolipoprotein B 60.71429 65.00000 29.83861 Negative correlation CE 18:3/total-c/HDL-c 67.24138 62.89308 −27.28733 LPC 18:2/apolipoprotein B 65.45455 60.12658 −16.19596 Total PC/apolipoprotein B 64.91228 61.39241 −12.91145 Total PC/total cholesterol 64.91228 60.75949 −10.60231 CE 16:1/HDL cholesterol 63.79310 61.63522 −22.01197 CE 18:3/HDL cholesterol 63.79310 62.26415 −24.04060 CE 18:3/LDL cholesterol 63.79310 62.26415 −27.21759 LPC 18:2/LDL-c/HDL-c 63.63636 61.39241 −21.70580 Total PC/total-c/HDL-c 63.15789 60.12658 −16.13350 PC 16:0/20:3/HDL cholesterol 62.50000 62.65823 −11.45558 PC 16:0/20:3/apolipoprotein B 62.50000 60.12658 −15.11848 CE 16:1/apolipoprotein B 62.06897 62.26415 −26.99397 CE 16:1/total cholesterol 62.06897 60.37736 −24.28770 CE 18:3/LDL-c/HDL-c 62.06897 62.89308 −31.33346 CE 18:3/apolipoprotein A-I 62.06897 60.37736 −23.09816 LPC 18:2/HDL cholesterol 61.81818 60.75949 −16.07209 LPC 18:2/apoA1/apoB 61.81818 63.92405 −19.23316 PC 16:0/20:4/LDL cholesterol 61.40351 60.12658 −15.98292 PC 16:0/20:4/apolipoprotein A-I 61.40351 60.75949 −12.23661 PC 16:0/20:4/total cholesterol 61.40351 61.39241 −13.37554 Total PC/LDL-c/HDL-c 61.40351 62.65823 −21.24583 PC 16:0/20:3/total-c/HDL-c 60.71429 61.39241 −19.07121 PC 18:0/20:4/apoA1/apoB 60.71429 62.02532 −13.36540 CE 18:3/apoA1/apoB 60.34483 60.37736 −27.55825 CE 18:3/apolipoprotein B 60.34483 61.00629 −25.31907 CE 20:5/HDL cholesterol 60.34483 60.37736 −23.45029 CE 20:5/LDL cholesterol 60.34483 60.37736 −25.93288 LPC 18:2/LDL cholesterol 60.00000 62.02532 −17.26480 LPC 18:2/total cholesterol 60.00000 62.65823 −14.92954 7b) Table of significant lipid to clinical ratios in LURIC study sorted by top sensitivity and specificity from subjects not undergoing statin treatment. Positive correlation Cer(d18:1/24:1)/apolipoprotein B 82.60870 62.06897 18.05222 Cer(d18:1/24:1)/total cholesterol 80.43478 61.20690 21.94547 LacCer(d18:1/20:0)/ 77.27273 67.00000 43.04967 apolipoprotein A-I Cer(d18:1/22:0)/LDL cholesterol 73.91304 60.34483 11.30545 Total Cer/LDL cholesterol 73.91304 62.39316 8.25598 Total Cer/total cholesterol 73.91304 70.08547 15.12131 LacCer(d18:1/20:0)/HDL cholesterol 72.72727 61.00000 39.32664 LacCer(d18:1/20:0)/apoA1/apoB 72.72727 60.00000 32.36387 Cer(d18:1/22:0)/total cholesterol 71.73913 61.20690 17.20335 Cer(d18:1/24:1)/LDL cholesterol 71.73913 61.20690 14.20759 Cer(d18:1/24:1)/total-c/HDL-c 71.73913 60.34483 13.34657 Total Cer/apolipoprotein A-I 71.73913 61.53846 19.09944 Total Cer/apolipoprotein B 71.73913 61.53846 12.34421 LacCer(d18:1/20:0)/LDL cholesterol 70.45455 61.00000 31.95659 LacCer(d18:1/20:0)/LDL-c/HDL-c 70.45455 61.00000 23.54484 Cer(d18:1/20:0)/LDL cholesterol 69.56522 60.34483 18.51094 Cer(d18:1/20:0)/apolipoprotein B 69.56522 68.10345 20.79615 Cer(d18:1/20:0)/total cholesterol 69.56522 71.55172 25.23403 Cer(d18:1/20:0)/total-c/HDL-c 69.56522 60.34483 14.96816 Cer(d18:1/22:0)/apolipoprotein B 69.56522 65.51724 14.20773 Cer(d18:1/24:0)/total cholesterol 69.56522 60.68376 11.30123 Cer(d18:1/24:1)/LDL-c/HDL-c 69.56522 62.93103 5.34922 Cer(d18:1/24:1)/apolipoprotein A-I 69.56522 60.34483 27.12684 Cer(d18:1/24:1)/triglycerides 69.56522 63.79310 32.27486 GlcCer(d18:1/24:1)/apolipoprotein B 69.56522 61.20690 17.03707 Total DAG/apolipoprotein A-I 68.88889 60.19417 15.46034 LacCer(d18:1/20:0)/total cholesterol 68.18182 61.00000 41.46919 PC O-34:1/apolipoprotein B 68.18182 60.00000 22.73127 Cer(d18:1/16:0)/LDL cholesterol 67.39130 62.06897 10.42412 Cer(d18:1/18:0)/total-c/HDL-c 67.39130 61.20690 21.75165 Cer(d18:1/18:0)/triglycerides 67.39130 64.65517 40.06643 Cer(d18:1/24:0)/LDL cholesterol 67.39130 62.39316 4.69599 Cer(d18:1/24:0)/apolipoprotein A-I 67.39130 61.53846 14.67208 GlcCer(d18:1/20:0)/total cholesterol 67.39130 60.34483 20.60005 Total GlcCer/apolipoprotein B 67.39130 60.34483 14.92180 Total LacCer/total cholesterol 67.39130 62.93103 23.69633 LacCer(d18:1/22:0)/apoA1/apoB 66.66667 61.76471 24.17423 LacCer(d18:1/22:0)/ 66.66667 60.78431 32.96039 apolipoprotein A-I LacCer(d18:1/20:0)/apolipoprotein B 65.90909 61.00000 37.03057 LacCer(d18:1/20:0)/total-c/HDL-c 65.90909 62.00000 33.48550 Cer(d18:1/16:0)/triglycerides 65.21739 62.06897 28.51323 Cer(d18:1/22:0)/apolipoprotein A-I 65.21739 66.37931 21.04730 Cer(d18:1/24:0)/apolipoprotein B 65.21739 62.39316 9.26738 GlcCer(d18:1/20:0)/apolipoprotein B 65.21739 60.34483 17.00627 GlcCer(d18:1/26:1)/ 65.21739 61.60714 21.90520 apolipoprotein A-I LacCer(d18:1/16:0)/triglycerides 65.21739 60.34483 25.01312 LacCer(d18:1/18:0)/apolipoprotein B 65.21739 61.40351 33.69302 LacCer(d18:1/18:0)/total cholesterol 65.21739 62.28070 38.87517 PS O-16:0/18:2-alkenyl/triglycerides 65.21739 61.60714 97.78843 PS O-16:1/18:2-alkyl/triglycerides 65.21739 61.60714 97.78843 Negative correlation CE 18:3/total-c/HDL-c 68.08511 62.28070 −27.28733 CE 18:3/LDL-c/HDL-c 63.82979 60.52632 −31.33346 CE 18:3/apoA1/apoB 61.70213 64.03509 −27.55825 CE 20:5/triglycerides 61.70213 60.52632 −15.26342 PC O-38:6/apolipoprotein A-I 60.86957 62.03704 −3.87776 Total LPC/apoA1/apoB 60.86957 60.71429 −14.81039 Total PC/apolipoprotein A-I 60.86957 62.50000 −8.50761 PC 18:0/20:4/apoA1/apoB 60.00000 64.28571 −13.36540

The preferred lipid molecules of the invention were selected as follows: a) it was likely to be biologically meaningful, b) it preferably belongs to a family of lipids that are behaving similarly, c) it is expressed in meaningful & measurable concentrations, d) it has very significant p-value or good AUC-value (>0.65) and for most also the %-change is substantial (>20%), and e) it appeared significant in different tests. About 15 lipids or lipid ratios, each with either a positive or negative CVD correlation, were selected based on the highest p-values and best sensitivity and specificity subjectively ensuring the balanced representation of all lipid classes. Sensitivity and specificity thresholds were annotated in cases where the threshold of 60 and 70 were reached, respectively. The preferred embodiment lipids, lipid-lipid ratios and lipid-clinical ratios are presented in tables 8-11.

TABLE 8 The preferred embodiment lipids selected from significant lipids detected from LURIC sample set. Percentage Lipid name p-value change Sensitivity Specificity Positive correlation Cer(d18:1/20:0) 0.00004 28.00357 LacCer(d18:1/20:0) 0.00010 32.91154 70.45455 60.00000 Cer(d18:1/24:1) 0.00039 23.37606 69.56522 60.34483 LacCer(d18:1/24:1) 0.00199 29.83279 PS O-18:2/16:0-alkenyl 0.00432 33.81177 PS O-16:1/18:2-alkyl 0.00590 32.17190 Total Cer 0.00932 15.44601 Total LacCer 0.01105 16.00541 GlcCer(d18:1/24:1) 12.75066 60.86957 62.06897 LacCer(d18:1/22:0) 0.00046 22.75541 Cer(d18:1/18:0) 0.00009 34.32550 Negative correlation Total PC 0.00921 −12.44220 PC 16:0/20:4 0.02256 −14.96966 Cer(d18:0/24:0) 0.03376 −21.87004 Total LPC 0.03443 −12.91576 CE 14:0 0.01090 −21.01258 CE 20:3 0.02157 −16.03606 CE 17:1 0.02204 −15.93952 PC 16:0/20:3 −18.78110 64.44444 60.71429 LPC 18:1 −14.45827 63.04348 60.71429 PC 18:0/20:3 0.00726 −17.20664 PC 18:0/18:1 0.00765 −18.18002 Cer(d18:0/22:0) 0.01158 −22.37263

TABLE 9 Preferred embodiments from significant lipid to lipid ratios detected from LURIC sample set. Lipid name/Lipid name p-value Percentage change Sensitivity Specificity Positive correlation GlcCer(d18:1/26:1)/Total CE 0.000000 35.356592 Cer(d18:1/24:1)/Total PC 0.000000 43.040800 Cer(d18:1/24:1)/PC 16:0/20:4 0.000000 59.876132 Cer(d18:1/20:0)/PC 16:0/20:4 0.000000 64.045788 LacCer(d18:1/20:0)/PC 16:0/20:3 0.000008 56.972652 80.39216 65.11628 Total Cer/Total PC 0.000011 29.949121 Total LacCer/Total PC 0.000012 37.815865 LacCer(d18:1/20:0)/PC 18:1/18:2 0.000013 48.432788 PS O-16:0/18:2-alkenyl/Total PS O 26.42621 97.82609 62.50000 Cer(d18:1/18:0)/PC 16:0/20:4 73.07370 80.00000 62.16216 LacCer(d18:1/20:0)/Total LPC 53.68111 78.84615 61.24031 LacCer(d18:1/20:0)/PC 16:0/20:4 82.44735 79.06977 68.42105 Negative correlation Cer(d18:0/24:0)/Cer(d18:1/24:1) 0.000003 −36.760563 Cer(d18:0/22:0)/Cer(d18:1/24:1) 0.000011 −34.086320 DAG 16:0/18:1/Total DAG −17.30528 88.88889 61.16505 Cer(d18:0/24:0)/Cer(d18:1/22:0) −32.40867 75.43860 66.25767 Cer(d18:0/24:0)/Total CE 0.0000000 −18.9531828 Cer(d18:0/24:0)/Cer(d18:1/24:1) 0.000003 −36.760563 Cer(d18:0/24:0)/Total Cer 0.0000049 −31.4531758 Cer(d18:0/24:0)/Cer(d18:1/18:0) 0.0000052 −40.2960344 Cer(d18:0/24:0)/PS O-16:0/18:2- 0.0000059 −40.8329837 alkenyl Cer(d18:0/24:0)/LacCer(d18:1/24:0) 0.0000067 −37.0287234 Cer(d18:0/22:0)/Cer(d18:1/18:0) 0.0000071 −36.5400374 Cer(d18:0/24:0)/Cer(d18:1/22:0) 0.0000105 −32.4086711 Cer(d18:0/22:0)/Cer(d18:1/20:0) 0.0000165 −31.5266149 Cer(d18:0/22:0)/PS O-16:0/18:2- 0.000045 −37.930990 alkenyl Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl 0.000045 −37.930990 GlcCer(d18:1/26:0)/LacCer(d18:1/20:0) −26.83109 80.48780 64.77273 Total LPC/Total LacCer 0.0000469 −26.08820 GlcCer(d18:1/26:0)/LacCer(d18:1/22:0) −21.27116 78.04878 61.79775

TABLE 10 Preferred embodiments from significant lipid to clinical ratios from LURIC sample set. Lipid name/Clinical measurement p-value Percentage change Sensitivity Specificity Positive correlation Cer(d18:1/20:0)/apolipoprotein A-I 0.00000 43.22923 Cer(d18:1/24:1)/apolipoprotein A-I 0.00000 36.15491 69.56522 60.34483 LacCer(d18:1/20:0)/apolipoprotein A-I 0.00001 47.52465 77.27273 67.00000 Total Cer/apolipoprotein A-I 0.00016 25.36811 Total LacCer/apolipoprotein A-I 0.00031 28.05057 71.73913 61.53846 Cer(d18:1/18:0)/apolipoprotein A-I 0.00001 49.43693 LacCer(d18:1/22:0)/apolipoprotein A-I 0.00001 35.92744 LacCer(d18:1/20:0)/HDL cholesterol 39.32664 72.72727 61.00000 Cer(d18:1/24:1)/apolipoprotein B 18.05222 82.60870 62.06897 Negative correlation Cer(d18:0/24:0)/apolipoprotein B 0.03428 −21.10483 Cer(d18:0/24:0)/total cholesterol 0.04309 −19.71324 Cer(d18:0/24:0)/apolipoprotein B 0.00181 −26.25271 PC 16:0/20:4/apolipoprotein B 0.00576 −15.39247 Cer(d18:0/24:0)/apolipoprotein A-I 0.01052 −22.39254

TABLE 11 Top candidates from each category, if available, are listed. The best candidates were selected based on following criteria: t-test p-value ≤0.05 and sensitivity ≥60% and specificity ≥60%. Measurement name p-value Percentage change Positive correlation Cer(d18:1/20:0) 0.00004 28.00357 LacCer(d18:1/20:0) 0.00010 32.91154 Cer(d18:1/24:1) 0.00039 23.37606 LacCer(d18:1/24:1) 0.00199 29.83279 LacCer(d18:1/22:0) 0.00046 22.75541 Cer(d18:1/18:0) 0.00009 34.32550 Cer(d18:1/24:1)/PC 16:0/20:4 0.000000 59.876132 LacCer(d18:1/20:0)/PC 16:0/20:3 0.000008 56.972652 PS O-16:0/18:2-alkenyl/Total PS O 26.42621 Cer(d18:1/18:0)/PC 16:0/20:4 0.000000 73.073704 Cer(d18:1/20:0)/apolipoprotein A-I 0.00000 43.22923 Cer(d18:1/24:1)/apolipoprotein A-I 0.00000 36.15491 LacCer(d18:1/20:0)/apolipoprotein A-I 0.00001 47.52465 Total LacCer/apolipoprotein A-I 0.00031 28.05057 LacCer(d18:1/20:0)/HDL cholesterol 39.32664 Cer(d18:1/18:0)/apolipoprotein A-I 0.00001 49.43693 Negative correlation PC 16:0/20:4 0.02256 −14.96966 Cer(d18:0/24:0) 0.03376 −21.87004 GlcCer(d18:1/26:0)/LacCer(d18:1/20:0) −26.83109 DAG 16:0/18:1/Total DAG −17.30528 Cer(d18:0/24:0)/Total Cer −31.45318 Total LPC/Total LacCer −26.08820 GlcCer(d18:1/26:0)/LacCer(d18:1/22:0) −21.27116 Cer(d18:0/24:0)/Cer(d18:1/24:1) 0.0000004 −36.1288066 Cer(d18:0/24:0)/Cer(d18:1/18:0) 0.0000052 −40.2960344 Cer(d18:0/24:0)/PS O-16:0/18:2-alkenyl 0.0000059 −40.8329837 Cer(d18:0/24:0)/Cer(d18:1/22:0) 0.0000105 −32.4086711 Cer(d18:0/22:0)/PS O-16:0/18:2-alkenyl 0.000045 −37.930990 Cer(d18:0/22:0)/PS O-16:1/18:2-alkyl 0.000045 −37.930990 Cer(d18:0/24:0)/LacCer(d18:1/24:0) 0.0000067 −37.0287234

Lipidomic analysis proved to be efficient in identifying novel plasma biomarkers for CVD complications.

Molecular lipid to molecular lipid ratio could be an important indicator of cellular lipid metabolism including e.g., enzyme activities in the lipid metabolism pathways. Thus, these ratios may provide more information as the absolute plasma concentrations of the molecular lipids alone. As the absolute molecular lipid plasma concentration differences in general between healthy individuals and atherosclerotic patients seem to be between 30-70%, it might be reasonable to calculate and use different ratios instead of absolute concentrations only. As lipoprotein particles (e.g. LDL, HDL, and VLDL) are serving as carriers for most of the lipids in the blood stream it is appropriate to relate molecular lipid concentrations to lipoprotein data. Thus, the molecular lipid to HDL-cholesterol, LDL-cholesterol, apolipoprotein A-I and apolipoprotein B ratios were calculated. In fact, a number of ratios between the concentrations of different molecular lipids outperformed absolute plasma concentrations as disease biomarkers in CVD patients.

As the detected lipids are carried in the lipoprotein particles (LDL, VLDL and HDL) it is obvious that the corresponding lipoprotein fraction concentrations will even improve the prediction potential of molecular lipids from the results of the present study in total serum/plasma samples.

The lipid lowering drug efficiency measurements have so far been based on LDL-C and HDL-C assays. As the inventors have herein observed more potential biomarkers that predict the development of high-risk CVD complications better than these classical analyses, future drug efficiency profiling should be based on new sensitive and specific biomarkers that are more directly related to the risk of severe CVD-related complications rather than to LDL-C.

Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, numerous equivalents to the specific embodiments described herein both in the Examples in the body of the entire patent description. Such equivalents are considered to be within the scope of this invention and are covered by the following claims. 

The invention claimed is:
 1. A method for determining whether a subject is at risk to develop one or more Cardiovascular Disease (CVD) complications, comprising: (a) assaying a sample from said subject to determine one or more lipid-lipid concentration ratio(s), wherein (an) increased or decreased lipid-lipid concentration ratio(s) in said sample, when compared to a control sample, is (are) indicative of said subject having an increased risk of developing one or more CVD complications, wherein the one or more lipid-lipid concentration ratio(s) whose increase(s) is (are) compared to the control is (are) selected from: Cer(d18:1/18:0)/PC 16:0/18:1, Cer(d18:1/20:0)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:2, Cer(d18:1/18:0)/PC 18:0/18:1, Cer(d18:1/20:0)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:2, Cer(d18:1/18:0)/PC 18:1/18:1, Cer(d18:1/20:0)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:2, Cer(d18:1/18:0)/PC 16:0/20:3, Cer(d18:1/20:0)/PC 16:0/20:3, Cer(d18:1/22:0)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 18:0/20:3, Cer(d18:1/24:1)/PC O-40:3, Cer(d18:1/26:0)/PC O-40:0, PC 16:0/16:0/PC 16:0/20:4, PC O-18:0/18:2-alkyl/PC O-36:5, PC O-32:0 (KDdiA-PC)/PC O-38:5, LacCer(d18:1/18:0)/PC 16:0/18:1, LacCer(d18:1/20:0)/PC 16:0/18:1, LacCer(d18:1/18:0)/PC 18:0/18:1, LacCer(d18:1/20:0)/PC 18:0/18:1, LacCer(d18:1/18:0)/PC 18:1/18:1, LacCer(d18:1/20:0)/PC 18:1/18:1, LacCer(d18:1/18:0)/PC 18:1/18:2, LacCer(d18:1/20:0)/PC 18:1/18:2, LacCer(d18:1/20:0)/PC 18:2/18:2, LacCer(d18:1/18:0)/PC 16:0/20:3, LacCer(d18:1/20:0)/PC 16:0/20:3, LacCer(d18:1/22:0)/PC 16:0/20:3, LacCer(d18:1/24:0)/PC 16:0/20:3, LacCer(d18:1/18:0)/PC 18:0/20:3, LacCer(d18:1/20:0)/PC 18:0/20:3, LacCer(d18:1/22:0)/PC 18:0/20:3, LacCer(d18:1/20:0)/PC 16:0/20:4, LacCer(d18:1/22:0)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 18:0/20:4, GlcCer(d18:1/20:0)/PC 16:0/20:4, Cer(d18:1/18:0)/SM (d18:1/17:2-OH), Cer(d18:1/18:0)/SM (d18:1/18:1), Cer(d18:1/18:0)/SM (d18:1/14:0) (d18:1/13:1-OH), Cer(d18:1/20:0)/SM (d18:1/14:0) (d18:1/13:1-OH), Cer(d18:1/24:1)/SM (d18:1/17:1-OH), Cer(d18:1/24:1)/SM (d18:1/18:0), Cer(d18:1/24:1)/SM (d18:1/23:0) (d18:1/22:1-OH), Cer(d18:1/24:1)/SM (d18:1/24:0) (d18:1/23:1-OH), LacCer(d18:1/20:0)/SM (d18:1/17:1-OH), LacCer(d18:1/20:0)/SM (d18:1/18:0) and LacCer(d18:1/22:0)/SM (d18:1/14:0) (d18:1/13:1-OH); and wherein the one or more lipid-lipid concentration ratio(s) whose decrease(s) is (are) compared to the control is (are) selected from: PC 18:1/18:2/PS O-16:0/18:2-alkenyl, PC 18:1/18:2/PS O-16:1/18:2-alkyl, PC 16:0/20:3/PS O-16:0/18:2-alkenyl, PC 16:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-16:0/18:2-alkenyl, PC 18:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-18:2/16:O-alkenyl, PC 16:0/20:4/PS O-16:0/18:2-alkenyl, PC 16:0/20:4/PS O-16:1/18:2-alkyl, PC 18:0/20:4/PS O-16:0/18:2-alkenyl, PC 18:0/20:4/PS O-16:1/18:2-alkyl, PC O-40:3/PS O-18:2/16:O-alkenyl, LPC 18:2/PS O-16:0/18:2-alkenyl, LPC 18:2/PS O-16:1/18:2-alkyl, LPC 16:0/LacCer(d18:1/20:0), LPC 16:0/LacCer(d18:1/22:0), LPC 16:0/LacCer(d18:1/24:0), LPC 16:0/LacCer(d18:1/24:1), LPC 18:1/LacCer(d18:1/20:0), LPC 18:2/LacCer(d18:1/20:0), Cer(d18:0/22:0)/LacCer(d18:1/24:0) and Cer(d18:0/24:0)/SM (d18:1/17:0) (d18:1/16:1-OH); or (b) assaying a sample from said subject to determine one or more lipid-clinical concentration ratio(s), wherein (an) increased or decreased lipid-clinical concentration ratio(s) in said sample, when compared to a control sample, is (are) indicative of said subject having an increased risk of developing one or more CVD complications, wherein the one or more lipid-clinical concentration ratio(s) whose increase(s) is (are) compared to the control is (are) selected from: Cer(d18:1/16:0)/total cholesterol, Cer(d18:1/18:0)/LDL cholesterol, PC O-32:0 (KDdiA-PC)/apolipoprotein A-I, PC O-32:0 (KDdiA-PC)/triglycerides, PC O-34:1/apolipoprotein B, PC O-34:1/triglycerides, LacCer(d18:1/18:0)/apolipoprotein A-I, LacCer(d18:1/18:0)/apolipoprotein B, LacCer(d18:1/18:0)/total cholesterol, LacCer(d18:1/18:0)/HDL cholesterol, LacCer(d18:1/18:0)/LDL cholesterol, LacCer(d18:1/18:0)/total-c/HDL-c, LacCer(d18:1/18:0)/triglycerides, LacCer(d18:1/20:0)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein B, LacCer(d18:1/20:0)/total cholesterol, LacCer(d18:1/20:0)/HDL cholesterol, LacCer(d18:1/20:0)/LDL cholesterol, LacCer(d18:1/20:0)/total-c/HDL-c, LacCer(d18:1/20:0)/LDL-c/HDL-c, LacCer(d18:1/20:0)/apoA1/apoB, LacCer(d18:1/20:0)/triglycerides, LacCer(d18:1/22:0)/apolipoprotein A-I, LacCer(d18:1/22:0)/apolipoprotein B, LacCer(d18:1/22:0)/total cholesterol, LacCer(d18:1/22:0)/HDL cholesterol, LacCer(d18:1/22:0)/LDL cholesterol, LacCer(d18:1/22:0)/total-c/HDL-c, LacCer(d18:1/22:0)/apoA1/apoB, LacCer(d18:1/22:0)/triglycerides, LacCer(d18:1/24:0)/apoA1/apoB, LacCer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/24:1)/apolipoprotein B, LacCer(d18:1/24:1)/total cholesterol and LacCer(d18:1/24:1)/triglycerides; and wherein the one or more lipid-clinical concentration ratio(s) whose decrease(s) is (are) compared to the control is (are) selected from: PC 18:0/18:1/LDL-c/HDL-c, PC 16:0/20:3/apolipoprotein B, PC 16:0/20:3/HDL cholesterol, PC 16:0/20:3/total-c/HDL-c, PC 16:0/20:3/LDL-c/HDL-c, PC 18:0/20:3/total-c/HDL-c, PC 18:0/20:3/LDL-c/HDL-c, PC 16:0/20:4/apolipoprotein A-I, PC 16:0/20:4/apolipoprotein B, PC 16:0/20:4/total cholesterol, PC 16:0/20:4/LDL cholesterol, PC 16:0/20:4/total-c/HDL-c, PC 16:0/20:4/LDL-c/HDL-c, PC 18:0/20:4/apoA1/apoB, PC O-38:6/apolipoprotein A-I, LPC 18:2/apolipoprotein B, LPC 18:2/total cholesterol, LPC 18:2/HDL cholesterol, LPC 18:2/LDL cholesterol, LPC 18:2/LDL-c/HDL-c and LPC 18:2/apoA1/apoB; wherein said CVD complications comprise myocardial infarction (MI) and/or death, and administering treatment to said identified CVD risk subject comprising administering a lipid modifying treatment that is a statin or any other HMG-CoA reductase inhibitor, niacin (nicotinic acid); a cholesterol absorption inhibitor; a cholesteryl ester transfer protein (CETP) inhibitor; a bile acids sequestrant; a fibrate; or a phytosterol.
 2. The method of claim 1, wherein (a) the one or more lipid-lipid concentration ratio(s) whose increase is compared to the control is selected from: Cer(d18:1/18:0)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:2, Cer(d18:1/18:0)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:2, Cer(d18:1/18:0)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:2, Cer(d18:1/18:0)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 18:0/20:3 and Cer(d18:1/24:1)/PC O-40:3; (b) the one or more lipid-lipid concentration ratio(s) whose decrease is compared to the control is selected from: PC 18:1/18:2/PS O-16:0/18:2-alkenyl, PC 18:1/18:2/PS O-16:1/18:2-alkyl, PC 16:0/20:3/PS O-16:0/18:2-alkenyl, PC 16:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-16:0/18:2-alkenyl, PC 18:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-18:2/16:O-alkenyl, PC 16:0/20:4/PS O-16:0/18:2-alkenyl, PC 16:0/20:4/PS O-16:1/18:2-alkyl, PC 18:0/20:4/PS O-16:0/18:2-alkenyl, PC 18:0/20:4/PS O-16:1/18:2-alkyl and PC O-40:3/PS O-18:2/16:O-alkenyl; (c) the one or more lipid-clinical concentration ratio(s) whose increase is compared to the control is selected from: Cer(d18:1/16:0)/total cholesterol, Cer(d18:1/18:0)/LDL cholesterol, PC O-32:0 (KDdiA-PC)/apolipoprotein A-I, PC O-32:0 (KDdiA-PC)/triglycerides, PC O-34:1/apolipoprotein B and PC O-34:1/triglycerides; and/or (d) the one or more lipid-clinical concentration ratio(s) whose decrease is compared to the control is selected from: PC 16:0/20:4/apolipoprotein A-I, PC 16:0/20:4/apolipoprotein B, PC 16:0/20:4/total cholesterol, PC 16:0/20:4/LDL cholesterol, PC 16:0/20:4/total-c/HDL-c, PC 16:0/20:4/LDL-c/HDL-c and PC 18:0/20:4/apoA1/apoB.
 3. The method of claim 1, wherein the one or more lipid-lipid concentration ratios comprises at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, or at least 8 lipid-lipid concentration ratios, and wherein the one or more lipid-clinical ratios comprises at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, or at least 8 lipid-clinical concentration ratios, or combinations thereof.
 4. The method of claim 1, wherein a) the CVD complication(s) is(are) atherosclerosis-induced; and/or b) the subject has atherosclerosis; or c) the subject does not have atherosclerosis.
 5. The method of claim 1, wherein a) the method further comprises determining the serum level of total cholesterol, low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), Apolipoprotein B (ApoB) and/or Apolipoprotein C-III (ApoC-III) in said sample; and/or b) the subject does not have elevated serum levels of one or more of total cholesterol, low-density lipoprotein cholesterol (LDL-C), Apolipoprotein C-III (ApoC-III) or Apolipoprotein B (ApoB), or a decreased serum level of HDL-cholesterol (HDL-C).
 6. The method of claim 1, wherein the subject a) is being or has been treated with one or more statins and/or any other HMG-CoA reductase inhibitor; or b) has not yet undergone statin therapy or therapy with any other HMG-CoA reductase inhibitor.
 7. The method of claim 1, wherein the lipid-lipid concentration ratio(s) or the lipid-clinical concentration ratio(s) is (are) assayed by using mass spectrometry, nuclear magnetic resonance spectroscopy, fluorescence spectroscopy or dual polarization interferometry, a high performance separation method, an immunoassay and/or with a binding moiety capable of specifically binding the analyte.
 8. The method of claim 6, wherein the statin is selected from the group consisting of atorvastatin, cerivastatin, fluvastatin, fluvastatin XL, lovastatin, pitavastatin, pravastatin, rosuvastatin and simvastatin.
 9. The method of claim 1, wherein the subject is at risk to develop or has suffered from one or more CVD complications.
 10. The method of claim 1, wherein the control sample is from (a) CAD patient(s) or a group of CAD patients that has/have remained free of any major CVD complications, wherein the sample is a blood sample, or a serum sample.
 11. The method of claim 1, wherein the sample is blood, serum or plasma and wherein the lipid-lipid concentration ratio(s) or the lipid-clinical concentration ratio(s) is (are) determined by using mass spectrometry.
 12. A method for evaluating the effectiveness of a lipid modifying treatment of CVD and/or one or more of its complications, in a subject, comprising: (a) assaying a sample from said subject to determine one or more lipid-lipid concentration ratio(s), wherein (an) increased or decreased lipid-lipid concentration ratio(s) in said sample, when compared to a control sample, is (are) indicative of effectiveness of said lipid modifying treatment, wherein the one or more lipid-lipid concentration ratio(s) whose decrease(s) is (are) compared to the control is (are) selected from: Cer(d18:1/18:0)/PC 16:0/18:1, Cer(d18:1/20:0)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:2, Cer(d18:1/18:0)/PC 18:0/18:1, Cer(d18:1/20:0)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:2, Cer(d18:1/18:0)/PC 18:1/18:1, Cer(d18:1/20:0)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:2, Cer(d18:1/18:0)/PC 16:0/20:3, Cer(d18:1/20:0)/PC 16:0/20:3, Cer(d18:1/22:0)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 18:0/20:3, Cer(d18:1/24:1)/PC O-40:3, Cer(d18:1/26:0)/PC O-40:0, PC 16:0/16:0/PC 16:0/20:4, PC O-18:0/18:2-alkyl/PC O-36:5, PC O-32:0 (KDdiA-PC)/PC O-38:5, LacCer(d18:1/18:0)/PC 16:0/18:1, LacCer(d18:1/20:0)/PC 16:0/18:1, LacCer(d18:1/18:0)/PC 18:0/18:1, LacCer(d18:1/20:0)/PC 18:0/18:1, LacCer(d18:1/18:0)/PC 18:1/18:1, LacCer(d18:1/20:0)/PC 18:1/18:1, LacCer(d18:1/18:0)/PC 18:1/18:2, LacCer(d18:1/20:0)/PC 18:1/18:2, LacCer(d18:1/20:0)/PC 18:2/18:2, LacCer(d18:1/18:0)/PC 16:0/20:3, LacCer(d18:1/20:0)/PC 16:0/20:3, LacCer(d18:1/22:0)/PC 16:0/20:3, LacCer(d18:1/24:0)/PC 16:0/20:3, LacCer(d18:1/18:0)/PC 18:0/20:3, LacCer(d18:1/20:0)/PC 18:0/20:3, LacCer(d18:1/22:0)/PC 18:0/20:3, LacCer(d18:1/20:0)/PC 16:0/20:4, LacCer(d18:1/22:0)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 18:0/20:4, GlcCer(d18:1/20:0)/PC 16:0/20:4, Cer(d18:1/18:0)/SM (d18:1/17:2-OH), Cer(d18:1/18:0)/SM (d18:1/18:1), Cer(d18:1/18:0)/SM (d18:1/14:0) (d18:1/13:1-OH), Cer(d18:1/20:0)/SM (d18:1/14:0) (d18:1/13:1-OH), Cer(d18:1/24:1)/SM (d18:1/17:1-OH), Cer(d18:1/24:1)/SM (d18:1/18:0), Cer(d18:1/24:1)/SM (d18:1/23:0) (d18:1/22:1-OH), Cer(d18:1/24:1)/SM (d18:1/24:0) (d18:1/23:1-OH), LacCer(d18:1/20:0)/SM (d18:1/17:1-OH), LacCer(d18:1/20:0)/SM (d18:1/18:0) and LacCer(d18:1/22:0)/SM (d18:1/14:0) (d18:1/13:1-OH); and wherein the one or more lipid-lipid concentration ratio(s) whose increase(s) is (are) compared to the control is (are) selected from: PC 18:1/18:2/PS O-16:0/18:2-alkenyl, PC 18:1/18:2/PS O-16:1/18:2-alkyl, PC 16:0/20:3/PS O-16:0/18:2-alkenyl, PC 16:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-16:0/18:2-alkenyl, PC 18:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-18:2/16:O-alkenyl, PC 16:0/20:4/PS O-16:0/18:2-alkenyl, PC 16:0/20:4/PS O-16:1/18:2-alkyl, PC 18:0/20:4/PS O-16:0/18:2-alkenyl, PC 18:0/20:4/PS O-16:1/18:2-alkyl, PC 0-40:3/PS O-18:2/16:O-alkenyl, LPC 18:2/PS O-16:0/18:2-alkenyl, LPC 18:2/PS O-16:1/18:2-alkyl, LPC 16:0/LacCer(d18:1/20:0), LPC 16:0/LacCer(d18:1/22:0), LPC 16:0/LacCer(d18:1/24:0), LPC 16:0/LacCer(d18:1/24:1), LPC 18:1/LacCer(d18:1/20:0), LPC 18:2/LacCer(d18:1/20:0), Cer(d18:0/22:0)/LacCer(d18:1/24:0) and Cer(d18:0/24:0)/SM (d18:1/17:0) (d18:1/16:1-OH); or (b) assaying a sample from said subject to determine one or more lipid-clinical concentration ratio(s), wherein (an) increased or decreased lipid-clinical concentration ratio(s) in said sample, when compared to a control sample, is (are) indicative of effectiveness of said lipid modifying treatment, wherein the one or more lipid-clinical concentration ratio(s) whose decrease(s) is (are) compared to the control is (are) selected from: Cer(d18:1/16:0)/total cholesterol, Cer(d18:1/18:0)/LDL cholesterol, PC O-32:0 (KDdiA-PC)/apolipoprotein A-I, PC O-32:0 (KDdiA-PC)/triglycerides, PC O-34:1/apolipoprotein B, PC O-34:1/triglycerides, LacCer(d18:1/18:0)/apolipoprotein A-I, LacCer(d18:1/18:0)/apolipoprotein B, LacCer(d18:1/18:0)/total cholesterol, LacCer(d18:1/18:0)/HDL cholesterol, LacCer(d18:1/18:0)/LDL cholesterol, LacCer(d18:1/18:0)/total-c/HDL-c, LacCer(d18:1/18:0)/triglycerides, LacCer(d18:1/20:0)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein B, LacCer(d18:1/20:0)/total cholesterol, LacCer(d18:1/20:0)/HDL cholesterol, LacCer(d18:1/20:0)/LDL cholesterol, LacCer(d18:1/20:0)/total-c/HDL-c, LacCer(d18:1/20:0)/LDL-c/HDL-c, LacCer(d18:1/20:0)/apoA1/apoB, LacCer(d18:1/20:0)/triglycerides, LacCer(d18:1/22:0)/apolipoprotein A-I, LacCer(d18:1/22:0)/apolipoprotein B, LacCer(d18:1/22:0)/total cholesterol, LacCer(d18:1/22:0)/HDL cholesterol, LacCer(d18:1/22:0)/LDL cholesterol, LacCer(d18:1/22:0)/total-c/HDL-c, LacCer(d18:1/22:0)/apoA1/apoB, LacCer(d18:1/22:0)/triglycerides, LacCer(d18:1/24:0)/apoA1/apoB, LacCer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/24:1)/apolipoprotein B, LacCer(d18:1/24:1)/total cholesterol and LacCer(d18:1/24:1)/triglycerides; and wherein the one or more lipid-clinical concentration ratio(s) whose increase(s) is (are) compared to the control is (are) selected from: PC 18:0/18:1/LDL-c/HDL-c, PC 16:0/20:3/apolipoprotein B, PC 16:0/20:3/HDL cholesterol, PC 16:0/20:3/total-c/HDL-c, PC 16:0/20:3/LDL-c/HDL-c, PC 18:0/20:3/total-c/HDL-c, PC 18:0/20:3/LDL-c/HDL-c, PC 16:0/20:4/apolipoprotein A-I, PC 16:0/20:4/apolipoprotein B, PC 16:0/20:4/total cholesterol, PC 16:0/20:4/LDL cholesterol, PC 16:0/20:4/total-c/HDL-c, PC 16:0/20:4/LDL-c/HDL-c, PC 18:0/20:4/apoA1/apoB, PC O-38:6/apolipoprotein A-I, LPC 18:2/apolipoprotein B, LPC 18:2/total cholesterol, LPC 18:2/HDL cholesterol, LPC 18:2/LDL cholesterol, LPC 18:2/LDL-c/HDL-c and LPC 18:2/apoA1/apoB; wherein said CVD complications comprise myocardial infarction (MI) and/or death, wherein said subject has been administered treatment comprising administering a lipid modifying treatment that is a statin or any other HMG-CoA reductase inhibitor, niacin (nicotinic acid); a cholesterol absorption inhibitor; a cholesteryl ester transfer protein (CETP) inhibitor; a bile acids sequestrant; a fibrate; or a phytosterol.
 13. The method of claim 1, wherein the treatment comprise administering a therapeutically effective dose of a drug capable of modulating one or more of the lipid-lipid concentration ratio(s) or lipid-clinical concentration ratio(s), wherein the one or more lipid-lipid concentration ratio(s) is (are) selected from: Cer(d18:1/18:0)/PC 16:0/18:1, Cer(d18:1/20:0)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:2, Cer(d18:1/18:0)/PC 18:0/18:1, Cer(d18:1/20:0)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:2, Cer(d18:1/18:0)/PC 18:1/18:1, Cer(d18:1/20:0)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:2, Cer(d18:1/18:0)/PC 16:0/20:3, Cer(d18:1/20:0)/PC 16:0/20:3, Cer(d18:1/22:0)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 18:0/20:3, Cer(d18:1/24:1)/PC O-40:3, Cer(d18:1/26:0)/PC O-40:0, PC 16:0/16:0/PC 16:0/20:4, PC O-18:0/18:2-alkyl/PC O-36:5, PC O-32:0 (KDdiA-PC)/PC O-38:5, LacCer(d18:1/18:0)/PC 16:0/18:1, LacCer(d18:1/20:0)/PC 16:0/18:1, LacCer(d18:1/18:0)/PC 18:0/18:1, LacCer(d18:1/20:0)/PC 18:0/18:1, LacCer(d18:1/18:0)/PC 18:1/18:1, LacCer(d18:1/20:0)/PC 18:1/18:1, LacCer(d18:1/18:0)/PC 18:1/18:2, LacCer(d18:1/20:0)/PC 18:1/18:2, LacCer(d18:1/20:0)/PC 18:2/18:2, LacCer(d18:1/18:0)/PC 16:0/20:3, LacCer(d18:1/20:0)/PC 16:0/20:3, LacCer(d18:1/22:0)/PC 16:0/20:3, LacCer(d18:1/24:0)/PC 16:0/20:3, LacCer(d18:1/18:0)/PC 18:0/20:3, LacCer(d18:1/20:0)/PC 18:0/20:3, LacCer(d18:1/22:0)/PC 18:0/20:3, LacCer(d18:1/20:0)/PC 16:0/20:4, LacCer(d8:1/22:0)/PC 16:0/20:4, LacCer(d18:1/20:0)/PC 18:0/20:4, GlcCer(d18:1/20:0)/PC 16:0/20:4, Cer(d18:1/18:0)/SM (d18:1/17:2-OH), Cer(d18:1/18:0)/SM (d18:1/18:1), Cer(d18:1/18:0)/SM (d18:1/14:0) (d18:1/13:1-OH), Cer(d18:1/20:0)/SM (d18:1/14:0) (d18:1/13:1-OH), Cer(d18:1/24:1)/SM (d18:1/17:1-0H), Cer(d18:1/24:1)/SM (d18:1/18:0), Cer(d18:1/24:1)/SM (d18:1/23:0) (d18:1/22:1-OH), Cer(d18:1/24:1)/SM (d18:1/24:0) (d18:1/23:1-OH), LacCer(d18:1/20:0)/SM (d18:1/17:1-OH), LacCer(d18:1/20:0)/SM (d18:1/18:0), LacCer(d18:1/22:0)/SM (d18:1/14:0) (d18:1/13:1-OH); PC 18:1/18:2/PS O-16:0/18:2-alkenyl, PC 18:1/18:2/PS O-16:1/18:2-alkyl, PC 16:0/20:3/PS O-16:0/18:2-alkenyl, PC 16:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-16:0/18:2-alkenyl, PC 18:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-18:2/16:O-alkenyl, PC 16:0/20:4/PS O-16:0/18:2-alkenyl, PC 16:0/20:4/PS O-16:1/18:2-alkyl, PC 18:0/20:4/PS O-16:0/18:2-alkenyl, PC 18:0/20:4/PS O-16:1/18:2-alkyl, PC O-40:3/PS O-18:2/16:O-alkenyl, LPC 18:2/PS O-16:0/18:2-alkenyl, LPC 18:2/PS O-16:1/18:2-alkyl, LPC 16:0/LacCer(d18:1/20:0), LPC 16:0/LacCer(d18:1/22:0), LPC 16:0/LacCer(d18:1/24:0), LPC 16:0/LacCer(d18:1/24:1), LPC 18:1/LacCer(d18:1/20:0), LPC 18:2/LacCer(d18:1/20:0), Cer(d18:0/22:0)/LacCer(d18:1/24:0) and Cer(d18:0/24:0)/SM (d18:1/17:0) (d18:1/16:1-OH); or wherein the one or more lipid-clinical concentration ratio(s) is (are) selected from: Cer(d18:1/16:0)/total cholesterol, Cer(d18:1/18:0)/LDL cholesterol, PC O-32:0 (KDdiA-PC)/apolipoprotein A-I, PC O-32:0 (KDdiA-PC)/triglycerides, PC O-34:1/apolipoprotein B, PC O-34:1/triglycerides, LacCer(d18:1/18:0)/apolipoprotein A-I, LacCer(d18:1/18:0)/apolipoprotein B, LacCer(d18:1/18:0)/total cholesterol, LacCer(d18:1/18:0)/HDL cholesterol, LacCer(d18:1/18:0)/LDL cholesterol, LacCer(d18:1/18:0)/total-c/HDL-c, LacCer(d18:1/18:0)/triglycerides, LacCer(d18:1/20:0)/apolipoprotein A-I, LacCer(d18:1/20:0)/apolipoprotein B, LacCer(d18:1/20:0)/total cholesterol, LacCer(d18:1/20:0)/HDL cholesterol, LacCer(d18:1/20:0)/LDL cholesterol, LacCer(d18:1/20:0)/total-c/HDL-c, LacCer(d18:1/20:0)/LDL-c/HDL-c, LacCer(d18:1/20:0)/apoA1/apoB, LacCer(d18:1/20:0)/triglycerides, LacCer(d18:1/22:0)/apolipoprotein A-I, LacCer(d18:1/22:0)/apolipoprotein B, LacCer(d18:1/22:0)/total cholesterol, LacCer(d18:1/22:0)/HDL cholesterol, LacCer(d18:1/22:0)/LDL cholesterol, LacCer(d18:1/22:0)/total-c/HDL-c, LacCer(d18:1/22:0)/apoA1/apoB, LacCer(d18:1/22:0)/triglycerides, LacCer(d18:1/24:0)/apoA1/apoB, LacCer(d18:1/24:1)/apolipoprotein A-I, LacCer(d18:1/24:1)/apolipoprotein B, LacCer(d18:1/24:1)/total cholesterol, LacCer(d18:1/24:1)/triglycerides; PC 18:0/18:1/LDL-c/HDL-c, PC 16:0/20:3/apolipoprotein B, PC 16:0/20:3/HDL cholesterol, PC 16:0/20:3/total-c/HDL-c, PC 16:0/20:3/LDL-c/HDL-c, PC 18:0/20:3/total-c/HDL-c, PC 18:0/20:3/LDL-c/HDL-c, PC 16:0/20:4/apolipoprotein A-I, PC 16:0/20:4/apolipoprotein B, PC 16:0/20:4/total cholesterol, PC 16:0/20:4/LDL cholesterol, PC 16:0/20:4/total-c/HDL-c, PC 16:0/20:4/LDL-c/HDL-c, PC 18:0/20:4/apoA1/apoB, PC O-38:6/apolipoprotein A-I, LPC 18:2/apolipoprotein B, LPC 18:2/total cholesterol, LPC 18:2/HDL cholesterol, LPC 18:2/LDL cholesterol, LPC 18:2/LDL-c/HDL-c and LPC 18:2/apoA1/apoB.
 14. The method of claim 13, wherein the drug is a statin or any other HMG-CoA reductase inhibitor, niacin (nicotinic acid); a cholesterol absorption inhibitor; a cholesteryl ester transfer protein (CETP) inhibitor; a bile acids sequestrant; a fibrate; or a phytosterol.
 15. The method of claim 14, wherein the statin is selected from the group consisting of atorvastatin, cerivastatin, fluvastatin, fluvastatin XL, lovastatin, pitavastatin, pravastatin, rosuvastatin and/or simvastatin.
 16. The method of claim 1, wherein the method further comprises: assaying a sample from said subject to determine the concentration(s) of one or more lipid(s), wherein (an) increased or decreased concentration(s) in said sample, when compared to a control sample, is (are) indicative of said subject having an increased risk of developing one or more CVD complications, wherein the one or more lipid(s) whose increase(s) in concentration is (are) compared to the control is (are) selected from: Cer(d18:1/16:0), Cer(d18:1/18:0), Cer(d18:1/20:0), Cer(d18:1/22:0), Cer(d18:1/24:1), Cer(d18:1/26:1), LacCer(d18:1/18:0), LacCer(d18:1/20:0), LacCer(d18:1/22:0), LacCer(d18:1/24:0), LacCer(d18:1/24:1), PS O-18:2/16:O-alkenyl, PS O-16:0/18:2-alkenyl, PS O-16:1/18:2-alkyl, PS O-18:0/18:2-alkenyl (PS O-18:1/18:2-alkyl) and PC O-32:0 (KDdiA-PC); and wherein the one or more lipid(s) whose decrease(s) in concentration is (are) compared to the control is (are) selected from: PC 18:0/20:4, PC 16:0/20:4, PC 18:0/20:3, PC 16:0/20:3, PC 16:0/22:6, PC 16:0/18:2, PC 18:0/18:1, PC 18:1/18:2, LPC 18:1, PC O-40:3, SM (d18:1/14:0) (d18:1/13:1-OH), SM (d18:1/23:0) (d18:1/22:1-OH), SM (d18:1/24:0) (d18:1/23:1-OH), Cer(d18:0/22:0) and Cer(d18:0/24:0).
 17. The method of claim 12, further comprising adjusting the treatment based on the assaying step of a) orb).
 18. The method of claim 12, wherein: (a) the one or more lipid-lipid concentration ratio(s) whose decrease is compared to the control is selected from: Cer(d18:1/18:0)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:1, Cer(d18:1/24:1)/PC 16:0/18:2, Cer(d18:1/18:0)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:1, Cer(d18:1/24:1)/PC 18:0/18:2, Cer(d18:1/18:0)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:1, Cer(d18:1/24:1)/PC 18:1/18:2, Cer(d18:1/18:0)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 16:0/20:3, Cer(d18:1/24:1)/PC 18:0/20:3 and Cer(d18:1/24:1)/PC O-40:3; (b) the one or more lipid-lipid concentration ratio(s) whose increase is compared to the control is selected from: PC 18:1/18:2/PS O-16:0/18:2-alkenyl, PC 18:1/18:2/PS O-16:1/18:2-alkyl, PC 16:0/20:3/PS O-16:0/18:2-alkenyl, PC 16:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-16:0/18:2-alkenyl, PC 18:0/20:3/PS O-16:1/18:2-alkyl, PC 18:0/20:3/PS O-18:2/16:O-alkenyl, PC 16:0/20:4/PS O-16:0/18:2-alkenyl, PC 16:0/20:4/PS O-16:1/18:2-alkyl, PC 18:0/20:4/PS O-16:0/18:2-alkenyl, PC 18:0/20:4/PS O-16:1/18:2-alkyl and PC O-40:3/PS O-18:2/16:O-alkenyl; (c) the one or more lipid-clinical concentration ratio(s) whose decrease is compared to the control is selected from: Cer(d18:1/16:0)/total cholesterol, Cer(d18:1/18:0)/LDL cholesterol, PC O-32:0 (KDdiA-PC)/apolipoprotein A-I, PC O-32:0 (KDdiA-PC)/triglycerides, PC O-34:1/apolipoprotein B and PC O-34:1/triglycerides; and/or (d) the one or more lipid-clinical concentration ratio(s) whose increase is compared to the control is selected from: PC 16:0/20:4/apolipoprotein A-I, PC 16:0/20:4/apolipoprotein B, PC 16:0/20:4/total cholesterol, PC 16:0/20:4/LDL cholesterol, PC 16:0/20:4/total-c/HDL-c, PC 16:0/20:4/LDL-c/HDL-c and PC 18:0/20:4/apoA1/apoB.
 19. The method of claim 12, wherein the method further comprises: assaying a sample from said subject to determine the concentration(s) of one or more lipid(s), wherein (an) increased or decreased concentration(s) in said sample, when compared to a control sample, is (are) indicative of effectiveness of said lipid modifying treatment, wherein the one or more lipid(s) whose decrease(s) in concentration is (are) compared to the control is (are) selected from: Cer(d18:1/16:0), Cer(d18:1/18:0), Cer(d18:1/20:0), Cer(d18:1/22:0), Cer(d18:1/24:1), Cer(d18:1/26:1), LacCer(d18:1/18:0), LacCer(d18:1/20:0), LacCer(d18:1/22:0), LacCer(d18:1/24:0), LacCer(d18:1/24:1), PS O-18:2/16:O-alkenyl, PS O-16:0/18:2-alkenyl, PS O-16:1/18:2-alkyl, PS O-18:0/18:2-alkenyl (PS O-18:1/18:2-alkyl) and PC O-32:0 (KDdiA-PC); and wherein the one or more lipid(s) whose increase(s) in concentration is (are) compared to the control is (are) selected from: PC 18:0/20:4, PC 16:0/20:4, PC 18:0/20:3, PC 16:0/20:3, PC 16:0/22:6, PC 16:0/18:2, PC 18:0/18:1, PC 18:1/18:2, LPC 18:1, PC O-40:3, SM (d18:1/14:0) (d18:1/13:1-OH), SM (d18:1/23:0) (d18:1/22:1-OH), SM (d18:1/24:0) (d18:1/23:1-OH), Cer(d18:0/22:0) and Cer(d18:0/24:0).
 20. A method for determining whether a subject is at risk to develop one or more Cardiovascular Disease (CVD) complications, comprising: assaying a sample from said subject to determine the concentration(s) of one or more lipid(s), wherein (an) increased or decreased concentration(s) in said sample, when compared to a control sample, is (are) indicative of said subject having an increased risk of developing one or more CVD complications, wherein the one or more lipid(s) whose increase(s) in concentration is (are) compared to the control is (are) selected from: Cer(d18:1/16:0), Cer(d18:1/18:0), Cer(d18:1/20:0), Cer(d18:1/22:0), Cer(d18:1/24:1), Cer(d18:1/26:1), LacCer(d18:1/18:0), LacCer(d18:1/20:0), LacCer(d18:1/22:0), LacCer(d18:1/24:0), LacCer(d18:1/24:1), PS O-18:2/16:O-alkenyl, PS O-16:0/18:2-alkenyl, PS O-16:1/18:2-alkyl, PS O-18:0/18:2-alkenyl (PS O-18:1/18:2-alkyl) and PC O-32:0 (KDdiA-PC); and wherein the one or more lipid(s) whose decrease(s) in concentration is (are) compared to the control is (are) selected from: PC 18:0/20:4, PC 16:0/20:4, PC 18:0/20:3, PC 16:0/20:3, PC 16:0/22:6, PC 16:0/18:2, PC 18:0/18:1, PC 18:1/18:2, LPC 18:1, PC O-40:3, SM (d18:1/14:0) (d18:1/13:1-OH), SM (d18:1/23:0) (d18:1/22:1-OH), SM (d18:1/24:0) (d18:1/23:1-OH), Cer(d18:0/22:0) and Cer(d18:0/24:0); and treating the subject indicated as being at risk of developing one or more CVD complications based on the assaying step, wherein the treating step comprises administering a lipid modifying treatment that is a statin or any other HMG-CoA reductase inhibitor, niacin (nicotinic acid); a cholesterol absorption inhibitor; a cholesteryl ester transfer protein (CETP) inhibitor; a bile acids sequestrant; a fibrate; or a phytosterol.
 21. The method of claim 20, wherein the CVD complications comprise myocardial infarction (MI) and/or death. 